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Molecular Biology

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Director’s Note:Overview:

HIMEDIA have been dedicated to customer-oriented innovative product development since its inception and are proudly serving the global scientific community since 1975. Over the years the company has diversified from being a Microbiology company to becoming a leader in Biosciences including Molecular Biology. The company product range includes from dehydrated culture media and bases, antibiotic differentiation discs, animal cell culture, plant tissue culture, molecular biology, high-purity chemicals and laboratory aids (plasticware, sophisticated instruments).

Mission:

At HiMedia our passion is to serve the scientific community worldwide and support innovation by developing highest quality products that provide cost-effective customer solutions. The scientific team at HiMedia take pride in providing customized solutions to research scientists and industry so that they can perform complex specialized experiments at no additional costs.

In this endeavour, the Molecular Biology Researchers at HiMedia are developing new tools for different molecular biology applications using highest purity Molecular Biology grade chemicals. These ultrapure chemicals are also used to manufacture density gradient separation media, kits to obtain ultrapure genetic material, rapid detection of pathogens using PCR, epigenetics and protein purification.

HiMedia’s mission is to continually provide solutions to scientists for most efficient sample processing, storage and transportation to ensure accurate molecular diagnosis. A dedicated technical team and product specialists in the molecular biology division at HiMedia are committed to provide on-field and customized technical solutions so that the researcher can focus on experimental design and results analysis.

Vision:

As a technology based Research and Development company, HiMedia are committed to simplifying sample processing and diagnosis to help the scientist improve specificity, sensitivity and efficiency of bioscience research. HiMedia have evolved immensely over the years to ensure highest customer satisfaction level.

Quality Objective:

HiMedia are ISO 9001:2008 accredited by BIS and DNV with ISO 13485, WHO GMP, USA FDA registered and CE Certification. HiMedia are proud to be associated with Strategic Partners covering all parts of India catering more than 20,000 customers across India, besides the coverage of more than 125 countries across the Globe. As a result, Himedia have been honored with the nation’s highest quality award “Rajiv Gandhi National Quality Award twice for the year 2011 and 2006, which re-confirms the commitment of HiMedia to develop quality system based Biosciences products in the service of humankind.

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INDEX

Sr. No. Title Page No

1. Introduction 3

2. DNA and RNA Extraction Kits 5

3. Buffers and Reagents for Electrophoresis 41

4. Molecular Biology Chemicals 43

5. Affinity Based Protein Purification 46

6. Protein Crystallography 46

7. Nucleo-Proteo Cards 47

8. Latex Agglutination Test Kits 48

9. Hematology Kits 49

10. Ladders/ Markers 50

11. Nucleic Acid Amplification 51

DNA Polymerase 52

Restriction Enzymes 56

Reagents and Master Mix 56

PCR Based Diagnostic Kits 59

12. Density Gradient Separation Media 63

13. Horizontal Electrophoresis Apparatus and Power Packs 65

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IntroductionSpin Column Based Nucleic Acid Extraction:

Spin columns contain a silica resin that selectively binds DNA/RNA, depending on the salt concentration and other factors influenced by the extraction method. Let us understand the principle of silica based column kits as follows:

Lysis:

Isolation of nucleic acids begins by lysis or breakdown of the cell wall. Destruction of protein structures is essential for release of nucleic acids from the nucleus. Lysis of soft tissue or cells is easy as compared to that of hard tissue (eg. plant material, bone, wood) which requires freezing in liquid nitrogen and subsequently pulverizing the tissue to a fine powder. The lysis formulas may vary based on whether you want DNA or RNA, but the common denominator is a lysis buffer containing a high concentration of chaotropic salt. Chaotropes destabilize hydrogen bonds, van der Waals forces, and hydrophobic interactions. Proteins are destabi-lized, including nucleases, and the association of nucleic acids with water is disrupted setting up the conditions for the binding to silica membrane.

Besides the chaotropes, there are usually some detergents involved to help with lysis, protein solubilization and pre-cipitation. There can also be enzymes used for lysis depending on the sample type. For eg. Proteinase K, (one such enzyme), works very well in such denaturing buffers; unlike lysozyme. Hence, lysozyme treatment is usually done before adding the denaturing salts.

Binding:

Column-based kits offer a more convenient protocol to isolate nucleic acids from samples. In short, the use of columns increases the throughput of samples, requires shorter time of isolation in comparison to traditional solvent based extraction, increases the yield of recovered nucleic acid, and improves the quality of purified nucleic acid suitable for downstream applications. Column based purification method either involves ion exchange or silica membrane technology.

When using silica membrane based purification, the presence of chaotropic salts or high salt concentration

is essential for denaturation of proteins which optimizes the selective binding of negatively charged nucleic acids to silica based membrane. In addition, to enhance and influence the binding of nucleic acids to silica membrane, alcohol is also added to the binding buffer. These conditions lead to an energetically favorable situation for nucleic acids (DNA / RNA) to adsorb to the silica membrane.

During the DNA / RNA adsorption step, unwanted primers, impurities like salts, enzymes, agarose, dyes, unincorporated nucleotides, detergents, oils and proteins will flow through without binding to the silica column.

Wash Steps:

Nucleic acids (DNA and RNA) are adsorbed to the silica surface, while majority of the other molecules pass through the column. However, the membrane is still contaminated with residual proteins and salts. The nucleic acids (DNA / RNA) bound to the silica column are then washed by applying appropriate buffer solutions to the column to remove any impurities, proteins and polysaccharides from the sample. The wash steps serve to remove these impurities. The first wash will often have a low amount of chaotropic salt to remove the protein and colored contaminants. This is always followed with an ethanol wash to remove the residual salts. The percent ethanol and its volume affects the yield of nucleic acid in the elution buffer, i.e. normally 80% alcohol will enable optimal binding of nucleic acids to silica membrane, while 70% or lower concentration of alcohol will dramatically reduce the binding. Too much alcohol will enhance binding of degraded nucleic acids and small species to the silica membrane. On the other hand, the use of too little (dilute) alcohol will reduce the efficiency of the wash buffer to wash away all of the salt from the membrane.

Dry Spin:

After the ethanol wash, most protocols have a centrifugation step to dry the column. This is to remove the ethanol and is essential for a clean eluate (DNA / RNA). Skipping the drying step may result in lower yields of nucleic acids. When the elution buffer (10 mM Tris buffer or molecular biology grade water) is applied to the membrane for elution, the nucleic acids become hydrated and are released

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from the membrane. In the presence of residual alcohol on the column, nucleic acids cannot be fully rehydrated.

Following are the indicators to detect ethanolcontamination:

1. Samples cannot be loaded in the agarose gel even in presence of loading dye.

2. Eluted samples do not freeze even when stored at -20oC.

Elution:

The final step is the release of pure nucleic acids (DNA / RNA) from the silica membrane. Elution efficiency isdependent on the pH and salt concentrations of the elution buffer. Elution is most efficient under alkaline conditions and low salt concentrations. The low salt concentrationallows renaturing of nucleic acids (DNA / RNA), causing them to lose affinity for silica membrane. For DNA

preparations, 10 mM Tris at a pH between 8-9 is used. DNA is more stable at a slightly alkaline pH and will dissolve faster in an alkaline buffer which is true even for DNApellets. The maximum elution efficiency is achievedbetween pH 7.0 and 8.5. Molecular Biology grade water tends to have a low pH, as low as 4-5 and high molecular weight DNA/RNA may not completely rehydrate in the short time used for elution. When using water to elute, ensure that the pH is within 7.0 - 8.5. In addition, elution of DNA/RNA can be enhanced by allowing the buffer to sit on the membrane for a few minutes beforecentrifugation. DNA/RNA eluted in water should be stored at -20oC to avoid degradation of DNA in the absence of a buffering agent. Elution with TE buffer (10mM Tris HCl, 1mM EDTA, pH 8.0) is not recommended as the presence of EDTA may inhibit subsequent enzymatic reactions such as PCR or restriction enzyme digestion.

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1) DNA-XPress™ Reagent (MB501)

Applications :

The DNA obtained is compatible with downstream applications such as

2) HiPurA™ Plant DNA Isolation Kit (CTAB Method) (MB502)Sample material : 300-400mg of plant tissue

Elution volume : 1 ml

Total DNA yield : Upto 500 μg (depending on plant type and species.)

A260 / A280 :1.6 - 1.9

Fig. 2 a : Lane 1 to 4: Coriander Leaf DNA

Fig. 1 a : Lane 1: Lambda DNA Hind III digest ladderLane 2, 3: Blood Genomic DNA

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Product Description :TM Reagent is a non–organic ready to use

reagent; based on Chomczynski method, which combines reliability and efficiency with the simplicity of a fast isolation protocol.

yeast and samples of viral origin can be used for isolation even from small or large volumes.

can inhibit PCR or other enzymatic reactions.

Principle / Technology :Solution based protocol Selective precipitation of DNA in Guanidine detergent lysis process.

Sample material : 1ml of blood, upto107 cells, 50-200mg of plant tissue, 50mg of animal tissue

Elution volume : Upto 300μl

Total DNA yield : Upto 50 μg

A260 / A280 : 1.6-1.9

Product Description :

presence of a rigid cell wall surrounding the plant cells.

used to break open plant cells and solubilize the contents.

from plant tissue in a chloroform/octanol step and this organic phase is separated by centrifugation. The top aqueous phase contains DNA and RNA.

precipitated and washed in organic solvents before re-dissolving in aqueous solution.

and fresh leaves.

The scale of extraction is dependent on the amount of starting material, for e.g. 300–400 mg of material requires 9 ml of Extraction Buffer and should yield upto 500 μg of DNA.

Principle / Technology : Solution based protocol :CTAB based extraction

DNA and RNA Extraction Kit

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Applications :


Product Description:

that irreversibly interact with proteins and nucleic acids during DNA isolation resulting in poor yield and purity of DNA.

for various downstream processes even from theHerbarium specimens.

compounds adhere to DNA in solution and form a colored extract around DNA, which can be removed by the addition of detergent.

co-precipitated and pure DNA is eluted in Elution Buffer.

Principle / Technology : Solution based protocol

Sample material : 100mg of plant tissue

Elution volume : 50 -100 μl

Total DNA yield : 7-12 μg from 100mg of wet tissue.

A260 / A280 : 1.6 - 1.9

3) HiPurA™Cotton DNA Isolation Kit (MB528)

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Fig. 2 b : Lane 1 & 2 : Wheat leaf DNA, Lane 3 : Hind III Marker

Applications


Fig. 3 a : Lane 1 to 3: Cotton leaf DNA

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4) HiPurA™ Silica Kit For DNA Isolation (MB503)

Applications :


Fig. 4 a : Lane 1 to 3: Recovered 1 kb band using MB503Lane 4: Reference 1 kb DNA ladder

1 2 3 4Product Description :

for extracting DNA from low melting agarose gels, chemically treated tissues, recovery of DNA from PCR, as well as for concentration of DNA without ethanol precipitation.

DNA is adsorbed on to glass powder particles.

buffer containing ethanol to remove excess of salt and impurities from sample.

Successive wash reduces salt concentration causing rehydration of the DNA leading to separation from glass powder

matrix. The clean DNA can be eluted in water or TE buffer.

than other organic solvent based extraction methods.


Sample material : Upto 400mg of agarose gel

Elution volume : 15-50μl

Total DNA yield : 70-80 % recovery.

A260 / A280 : 1.6 - 1.9

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Product description:

used to break open plant cells and solubilize the contents.

plant tissue in an organic chloroform-isoamylalcohol step and separated by centrifugation.

removed by the addition of RNase A, the DNA is precipitated and washed in organic solvents before redissolving inaqueous solution.

The scale of extraction is dependent on the amount of starting material. The sample used can be fresh or freeze dried.


Sample material : 200mg of plant tissue

Elution volume : 100 μl

Total DNA yield : 10-50 μg from 200mg of wet tissue.

A260 / A280 : 1.6 - 1.9

Applications :


5) HiPurATM Banana DNA Isolation Kit (MB533)

Fig. 5 a : Lane 1 & 2: Banana leaf DNA

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Fig. 6 a : Lane 1to 3: Human Blood Genomic DNA

Product Description :

The kit simplifies isolation and purification of high molecular weight genomic DNA from fresh, frozen and anticoagulated whole blood by using highly efficient solution based system.

from bone marrow, buffy coat and cultured cells.

of red blood cells followed by lysis of white blood cells and their nuclei.

and short washing steps while high molecular weightgenomic DNA remains in the solution. High quality genomic DNA is then purified by isopropanol precipitation.

DNA. This system can be scaled up from 0.1 ml to 12 ml of whole blood in a single tube.

inhibitors from DNA for reliable downstream applications.


Sample material :0.1ml to 12ml whole blood

Elution volume :Elute in 100μl-1 ml of warm Elution buffer depending upon the volume of starting material

Total DNA yield :Upto 700 μg

A260 / A280 :1.6 - 1.9

6) HiPurATM SPP Blood DNA Isolation Kit (MB541)

Applications :


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Fig. 7 a : Lane 1 to 4 : Human Blood Genomic DNA using HiPurA™ Blood Genomic DNA Maxiprep Purification Kit (MB517)

Fig. 7 b : Lane 1 to 3 : Human Blood DNA using HiPurA™ Blood Genomic DNA Miniprep Purification Kit (MB504)

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silica-membrane-based DNA purification from fresh, old (more than 24 hours) and frozen blood.

incubation of whole blood in a solution containing chaotropic ions in the presence of Proteinase K at 55oC.

spin column, and impurities like proteins, polysaccharides, low molecular weight metabolites and salts are removed by short washing steps.

multiple number of samples in various volumes. The columns have a high binding capacity and high quality DNA is obtained which is suitable for downstream processes.

as well as vaccum manifold processes.

blood samples.

binding properties of the advanced gel membrane and the speed plus versatility of vacuum technology to yield high quantity of DNA.

Principle / Technology : Silica column based protocol

Formats available : Mini Midi Maxi 96-Wells Upto

Total blood volume : 200μl 1-2 ml 10ml 200μl

Elution volume : 200μl 300μl 2 ml 100μl

Total DNA yield :. 4-12μg 15-50μg 100-600μg 4-12 μg

A260 / A280 : 1.6 - 1.9

7) HiPurA™ Blood Genomic DNA Miniprep Purification Kit (MB504) HiPurA™ Blood Genomic DNA Midiprep Purification Kit (MB516) HiPurA™ Blood Genomic DNA Maxiprep Purification Kit (MB517) HiPurA™ 96 Blood Genomic DNA Purification Kit (MB521)

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Applications:

The purified DNA can be used in a wide range of downstream applications including:

Fig. 7 c : Lane 1 & 2 : DNA from Frozen Blood Lane 3 & 4 : DNA from normal blood using

HiPurA™ Blood Genomic DNA Miniprep Purification Kit (MB504)

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provides a convenient method for extracting DNA from bacteria (Gram positive and Gram negative).

high binding capacity resulting into best quality DNA from various bacterial species.

peptidoglycan cell wall as compared to Gram negative bacteria.These cell walls are lysed efficiently by using

Proteinase K unlike Gram positive cell wall which are lysed completely using lysozyme and Proteinase K in combination.

with centrifugation as well as vacuum manifold processes.

bacterial species.


Formats available : miniprep 96 well format

Sample material : 1.5 ml culture 1.5ml culture

Elution volume : 200 μl 200 μl

Total DNA yield :. upto 20 μg upto 20 μg

A260 / A280 :1.6 - 1.9

8) HiPurATM Bacterial Genomic DNA Purification Kit (MB505) HiPurATM 96 Bacterial Genomic DNA Purification Kit (MB548)

Fig. 8 b : Lanes 1 to 12, 13 to 24, 25 to 36 : shows representative gel picture of bacterial genomic DNA extracted using HiPurATM 96 Bacterial Genomic DNA Purification Kit (MB548) from different bacterial species.

Fig. 8 a : Lanes 1 to 3 :DNA from E. coliLanes 4 to 6 :DNA from S. aureus using

HiPurATM Bacterial Genomic DNA Purification Kit (MB505)

Applications:

The purified DNA can be used in a wide range of downstream applications including:

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13 14 15 16 17 18 19 20 21 22 23 24

25 26 27 28 29 30 31 32 33 34 35 36

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cellulose and polysaccharides cell wall.

physical aberration of the tissue i.e. grinding plant materialin liquid nitrogen along with lysis buffer.

contaminants using a shredder. The flow through fraction is mixed with a binding solution to ensure efficient binding of DNA to the column.

obtain high quality DNA in the elution buffer.

plants like rice, sorghum, mint coriander, Okra, Ashwagandha, Eucalyptus, etc.

with centrifugation as well as vacuum manifold processes.

bacterial species.


Formats available : Mini prep Maxi prep 96-well format

Sample material : 100mg 1g 50-100mg

Elution volume : 200 μl 1ml 200μl

Total DNA yield :. 5-40μg 50-250 μg 1-15μg

A260 / A280 :1.60 - 1.90

9) HiPurATM Plant Genomic DNA Miniprep Purification Kit (MB507) HiPurATM Plant Genomic DNA Maxiprep Purification Kit (MB520) HiPurATM 96 Plant Genomic DNA Purification Kit (MB523)

Fig. 9 a : 1 - Mint leaf DNA, 2 - Ashwagandha leaf DNA 3 - Eucalyptus leaf DNA

Fig. 9 b : Lane 1 : Hind III ladder,Lane 2 & 3 : Castor seed DNA,Lane 4 & 5 : Groundnut DNA

Fig. 9 c : 4 - Sorghum DNA, 5 - Rice leaf DNA, 6 - Wheat leaf DNA

Oil & Medicinal Plant DNA

Monocot Plant DNA

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Fig. 9 e : 9 : Sunflower leaf DNA 10 : Tomato leaf DNA

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Fig. 9 f : 11 : Arabidopsis leaf DNA 12 :13 : Lane 1: Hind III ladder Lane 2 to 4 : Murraya (Curry leaf) leaf DNA14 : Lane 1 to 3 : Cabbage leaf DNA

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12

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Fig. 9 d : 6 : Bamboo leaf DNA7 : Sugarcane leaf DNA8 : (Conifer) Pine leaf DNA

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Other Plant DNA

Applications:

The purified DNA is suitable for downstream applications, such as:

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Plasmid purification follows the principle of advanced silica based membrane technology in the form of spin column.

membrane, washing of residual contaminants and elution of clean plasmid DNA.

neutralization helps in elimination of the genomic DNA but conserves the plasmid DNA.

plasmid, but yields may vary depending on type of plasmid.

volumes of culture.

as well as vacuum manifold processes.

bacterial species.

Principle / Technology :Silica column based protocol

Formats available : Mini Midi Maxi 96-well prep prep prep format

Sample material : 1-5 ml 150 ml 500 ml 1-2ml

Elution volume : 50μl 2ml 3ml 150μl

Total DNA yield: up 20μg 25 μg- upto 20 μg from 100 μg 500 μg 1-2 ml of

E.coli culture

A260 / A280 : 1.6 .1.9

10) HiPurATM Plasmid DNA Miniprep Purification Kit (MB508) HiPurATM Plasmid DNA Midiprep Purification Kit (MB509) HiPurATM Plasmid DNA Maxiprep Purification Kit (MB510) HiPurA™ 96 Plasmid DNA Purification Kit (MB518)

Fig. 10 b : Different Plasmid DNA Purified using MB508

Lanes Plasmid Plasmid Yield A260/ A260/

Name size (ng/μl) A280 A230

1 1 kb DNA Ladder

2 pUCGFP 2-2.5kb 117.1 1.81 2.00

3 E3 Prima 2-2.5kb 77.0 1.84 2.16

4 pGFPN1 3kb 71.6 1.83 2.37

5 pTZ257R 6-8kb 39.4 1.76 2.12

6 pEG202 6-8kb 63.6 1.90 2.37

8 pUC19 1.5-2kb 28.9 1.84 2.87

9 p3939 4kb 50.4 1.83 2.59

10 Hind III Marker

Applications:

Plasmid Miniprep Purification Kit provide high purity DNA suitable for use in most applications including:

In vitro transcription & translation

1 2 3 4 5 6 7 8 9 10

Fig. 10 a : Lane 1 to 4 : Plasmid DNA Purified using HiPurATM Plasmid DNA Midiprep Purification Kit (MB509)

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DNA from PCR & other enzymatic reactions. Fragments ranging from 100 bp to 10kb can be purified from primers, nucleotides, polymerases & salts by the use of silica membrane.

(up to 50 bp), proteins, MgCl2, mineral oil etc are completely removed in the wash step.

of low salt buffer. The PCR product recovery is 80-95% depending on the size of DNA.

the time required to purify large number of PCR products obtained from various simultaneously amplified DNA samples. This kit is compatible to vacuum manifold and centrifuge.


Formats available : Miniprep 96-well format

Sample material : 25-50 μl of 25-50 μl of PCRPCR reaction reaction

Elution volume : 30 - 50μl 150μl

Total DNA yield : 80-90% recovery 80-90% recovery

A260 / A280 : 1.6 - 1.9

Applications:

DNA fragments purified are ready to use in downstreamapplications including:

11) HiPurATM PCR Product Purification Kit (MB512)HiPurA™ 96 PCR Product Purification Kit (MB519)

Fig. 11 a : Lane 1 : 100 bp DNA ladder Lane 2 : PCR product before purification

(primer-dimers are marked in red circle)Lane 3, 4 : PCR product after purification

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on the cell membrane of Gram negative bacteria such as E.coli. During the lysis step of plasmid purification, endotoxinsare released, which significantly reduce transfection efficiencies in endotoxin sensitive cell lines.

transfection experiment setup, influencing the outcome and reproducibility of results and making them difficult to compare and interpret. In gene therapy research, endotoxins can interfere by causing endotoxic-shock syndrome and activation of the complement cascade.

TM Endotoxin free Plasmid Purification Kit integrates an efficient endotoxin removal step into plasmid purification procedure. Cells are lysed and endotoxins are selectively precipitated in a proprietary buffer.

for binding of the DNA molecules in the presence of high salt concentration. The adsorbed DNA is washed to remove contaminants and the pure plasmid DNA is eluted in elution buffer.

EU/μg of DNA This kit gives high yields for high- as well as lowcopy number plasmids.

Principle / Technology : Silica spin column based protocol

Formats available : Mini prep, Midi prep, Maxi prep

Sample material : 1-5 ml 150 ml 500 ml

Elution volume : 50 μl 2 ml 3 ml

Total DNA yield : upto 20 μg 25μg -100μg 500 μg(high & low (high & low (high & lowcopy) copy) copy)

A260 / A280 : 1.60 - 1.90

12)HiPurATM Endotoxin free Plasmid DNA Miniprep Purification Kit (MB513) HiPurATM Endotoxin free Plasmid DNA Midiprep Purification Kit (MB514) HiPurATM Endotoxin free Plasmid DNA Maxiprep Purification Kit(MB515)

Applications:

Endo Free Plasmid kits yield endotoxin free Plasmid DNA which is well suited for sensitive applications including

Fig. 12 a : Lane 1 : 1kb ladder Lane 2 & 3 : pUCGFP plasmid DNA extracted using

HiPurATM Endotoxin free Plasmid DNA Maxiprep Purification Kit (MB515)

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Product description:TM Mammalian Genomic DNA Purification Kit provides

a convenient method for extracting DNA from animal tissue (spliced and digested), suspended/attached animal cells, and whole blood by the spin-column procedure.

the contaminants of cell matrix and non nucleic acid component of cells.

liquid nitrogen is advised if tissue is preserved or hard.


Formats available : Miniprep

Sample material :1 x 106 cells from culture cells, upto 25mg of tissue

Elution volume : 200 μl

Total DNA yield : 10-20 μg from 1 x 106 cells, (30-45 g from upto 25mg oftissue yield can change depending on cell line and tissue)

A260 / A280 : 1.6 - 1.9

13) HiPurA™ Mammalian Genomic DNA Purification Kit (MB506)

Fig. 13 a : Lane 1: 1 kb DNA ladderLane 2, 3: Mammalian Genomic DNA

Applications:

The purified DNA can be used in a wide range of downstream applications including

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cells and sperm cells. HiPurATM Sperm Genomic DNA Purification Kit simplifies selective isolation of DNA from sperm cells out of this mixture.

containing DL-Dithiothreitol (DTT) eliminates thecontamination of the non-sperm cell DNA.

DNA which is washed in two rapid steps to remove trace salt and protein contaminants resulting in the elution of high quality DNA.

samples as well.



Sample material :100-200 μl semen sample.

Elution volume : 200μl

Total DNA yield :2μg-15μg

14) HiPurA™ Sperm Genomic DNA Purification Kit (MB522)

Fig. 14 a : Lane 1: Lambda DNA HindIII digest ladderLane 2, 3: Sperm genomic DNA

A260 / A280 : 1.6 - 1.9

Applications

The DNA obtained using HiPurATM Sperm Genomic DNA Miniprep Purification Kit is compatible with the following downstream applications

Product description:TM Forensic Sample Genomic DNA Purification Kit

simplifies isolation of DNA from cigarette butts, hair, nail clippings, blood, saliva or semen stains with spin-column procedure.

column comprises of three steps i.e. adsorption of DNA to the membrane, removal of residual contaminants and elution of pure genomic DNA.

TM

Miniprep Spin Column to yield high purity DNA. Two rapid wash steps remove trace salt and protein contaminants resulting in the elution of high quality DNA in the Elution Buffer provided with the kit.

Principle / Technology : Silica spin column based protocol


Sample material : 0.5-2.5 cm pieces of cigarette butt, hair, nail clipping.

15) HiPurA™ Forensic Sample Genomic DNA Purification Kit (MB524)1 2 3

Fig. 15 a : Lane 1 : Hind III Marker, Lane 2 & 3 : DNA from semen stained sample

1 2 3

Elution volume : 20- 50μl

Total DNA yield : 2μg-15μg

A260 / A280 : 1.70 - 1.90

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tool for the identification of missing individuals & unknown remains.

isolation difficult. Hence, HiPurATM Bone DNA Purification Kit allows simplified DNA isolation using Bone Extraction Buffer for decalcification.

powder suspension with the help of binding buffer under optimal conditions.

various downstream application.



Sample material : 500mg


A260 / A280 : 1.6 - 1.9

16) HiPurA™ Bone DNA Purification Kit (MB525)

Fig. 16 a : Lane 1, 2 : DNA from chicken boneLane 3 : Lambda DNA HindIII digest ladder

Applications :

The DNA obtained using HiPurATM Bone DNA Purification Kit is compatible with the following downstream applications

1 2 3

Fig. 15 b : PCR for 18S rRNA using DNA Extracted from various forensic sample.

Lane 1,2,3 : DNA from semen sample Lane 4,5,6 : DNA from blood stained clothLane 7,8,9 : DNA from blood stained paperLane 10,11 : DNA from Hair roots Lane 12,13 : DNA from NailsLane 14 : 1kb DNA Ladder

Applications :1 2 3 4 5 6 7 8 9 10 11 12 13 14

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Product description:TM Streptomyces DNA Purification Kit simplifies

purification of DNA from Streptomyces spp. grown in log phase.

by Proteinase K digestion in combination with lysis buffer.

followed by two rapid wash steps to yield high quality purified DNA.



Sample material : 10-15mg wet weight



A260 / A280 : 1.6 - 1.9

17) HiPurA™ Streptomyces DNA Purification Kit (MB527)

Fig. 17 a : Lane 1 & 2 : DNA from Streptomyces griseusLane 3 & 4 : DNA from Streptomyces laverndulae

Applications:

The DNA obtained using HiPurATM Streptomyces DNA Purification Kit is compatible with the following downstream applications

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Fig. 18 a : Lane 1 to 2 : Insect (Housefly) DNA Lane 3 : Hind III Marker

Product description:TM Insect DNA Purification Kit provides

a fast and easy method for purification of total DNA from insects, arthropods, roundworms, flatworms etc. (fresh or frozen samples).

lysis by Proteinase K in a chaotropic salt solution.

Following lysis, DNA is bound to the silica membrane.Subsequent wash steps eliminates trace salts & othercontaminants to yield highly purified DNA.



Sample material : 50mg wet weight



A260 / A280 : 1.6 - 1.9

18) HiPurATM Insect DNA Purification Kit (MB529)

Applications :

The DNA obtained using HiPurATM Insect DNA Purification Kit is compatible with the following downstream applications

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HiPurATM Paraffin-Embedded Tissue DNA Purification Kit simplifies isolation of DNA from FFPE tissues such as liver, brain, lung, heart, kidney, spleen etc.

sample preservation and archiving. These archived samples could potentially provide a wealth of information in retrospective molecular studies of diseased tissues.

is subjected to organic solvent treatment to eliminate traces of paraffin.

chaotropic salt solution. Following lysis is the binding of DNA to the silica gel membrane to yield high purity DNA.

contaminants resulting in the elution of high quality DNA in the elution Buffer provided with the kit.



Sample material :25 mg paraffin embedded tissue



A260 / A280 : 1.6 - 1.9

19) HiPurATM Paraffin-Embedded Tissue DNA Purification Kit (MB530)

Applications:

The DNA obtained using HiPurATM Paraffin-Embedded Tissue DNA Purification Kit is compatible with the following down stream applications

Fig. 19 a : Lane 1 : Hind III Marker Lane 2 to 3 : Paraffin-Embedded Tissue DNA

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Product description :

Isolation of the buccal DNA is mostly preferred for the studiesof the conserved sequences and the homo and heterozygosity of the allelic studies.

TM Buccal DNA Purification Kit simplifies isolation of DNA from the inside of cheek. The DNA purificationprocedure using the miniprep spin column comprises of three steps i.e. adsorption of DNA to the membrane, removal of residual contaminants and elution of pure genomic DNA.

high purity of DNA. Two rapid wash steps remove trace salts and protein contaminants resulting in the elution of high quality DNA in the Elution Buffer provided with the kit.




Total DNA yield : 0.5μg-3.5μg

A260 / A280 : 1.6 - 1.9

20) HiPurATM Buccal DNA Purification Kit (MB531)

Applications :

The DNA obtained using HiPurATM Buccal DNA Purification Kit is compatible with the following downstream applications

Fig. 20 a : Lane 1 to 4 : Extracted buccal DNA Lane 5 : 1 kb reference ladder

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Product description : TM Quick Gel Purification Kit simplifies isolation of

DNA from agarose gel using silica binding principle.

57°C in a gel binding buffer, that helps DNA to bind to silica membrane. The adsorbed DNA is washed to remove salt and impurities from the original sample and the clean DNA is eluted in Elution Buffer.

the pH shift, since pH of the solution determines the nucleic acid binding efficiency of the silica membrane.

completely removed in wash step, leaving pure nucleic acid to be eluted in the elution buffer provided with the kit. The purified DNA can be used for further downstream applications.

grades of agarose gel giving upto 90% recovery of ready to use DNA.



Sample material : Upto 400mg of gel with DNA per prep.

Elution volume : 30 - 50μl

Total DNA yield : 80-90% recovery

A260 / A280 : 1.6 - 1.9

21) HiPurATM Quick Gel Purification Kit (MB539)

Fig. 21 a : Lane 1 to 5 : Gel extracted 1 kb Band Lane 6 : 1 kb ladder

Applications:

DNA fragments purified are ready to use directly in downstream applications, including:

in vitro transcription

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Product description :

parameters like organic matter contents, pH, humic acid content, metal components etc. These compounds render difficulties in DNA extraction or result in co-elution of the inhibitors along with the DNA.

downstream processes like PCR, restriction digestion.TM Soil DNA Purification Kit provides a fast and easy

method for purification of total DNA from environmental samples containing a high humic acid content including difficult soil types such as compost, sediment, manure and other common soil types.

is effective in removing PCR inhibitors from even the most difficult soil types.

step. The total genomic DNA obtained is bound to silica membrane of column, washed and eluted from the membrane.



Sample material : 250-500 mg


Total DNA yield :Upto 20μg

A260 / A280 :1.6 - 1.9

22) HiPurATM Soil DNA Purification Kit (MB542)

Applications:

The DNA obtained using HiPurATM Soil DNA Purification Kit is compatible with the following downstream applications

1 2 3 1 2 3 44

1 2 3 4 5

1 2 3 4 5

1Fig. 22 a : 1 :

Lane 4 : Hind III Marker

2 : Lane 1 to 3 : DNA from Geranium compost soil Lane 4 : Hind III Marker

Fig. 22 b : 1 : Lane 1 & 2 : DNA from Clayey soil Lane 3 & 4 : DNA from Sandy soil Lane 5 : Hind III Marker

Fig. 22 b : 2 : 16S r RNA PCR using extracted DNA formsoil samples

Lane 1 & 2 : Amplified product from Clayey soilLane 3 & 4 : Amplified product from Sandy soil Lane 5 : 100 bp ladder

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Molecular Biology

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Fig. 23 a : DNA extracted from various fungal cultures.Lane 1 to 3 : Trichoderma spp. DNA Lane 4 to 6 : Aspergillus niger DNALane 7 to 9 : Penicillium notatum DNA


them difficult to lyse. Therefore, they have to be lysedusing mechanical force to rupture the tissue.

DNA from fungal cells.TM Fungal DNA Purification Kit simplifies the

isolation of DNA from fresh fungal material with spin column procedure.

The sample is ground in liquid nitrogen along with Lysis Buffer. Protein precipitation is followed by removal of other contaminants using HiShredder. The flow-through fraction is then mixed with a solution that enhances the binding of DNA to the column.

This solution is then passed through silica column that isfollowed by washing steps to remove trace contaminants.High quality DNA is eluted in the elution Buffer.



Sample material : 100-150 mg of wet weight.


Total DNA yield : 2μg-30μg (depending on species)

A260 / A280 : 1.6 - 1.9

23) HiPurATM Fungal DNA Purification Kit (MB543)

Applications:

The DNA obtained using HiPurA Fungal DNA Purification Kit is compatible with the following downstream applications

1 2 3 4 5 6 7 8 9

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epithelium, bacteria, parasites if present, which can be the source of DNA .

Stool DNA can be used as the means of diagnostic test for detection of the specific bacteria, fungus and parasitic pathogens with PCR for human clinical diagnosis or particular wild lifestudies etc.

Inhibitor Removal Solution (IRSH) provided in the kit eliminates the inhibitors, yielding high quality DNA suitable for all downstream applications.



Sample material : 250 mg stool sample.


Total DNA yield : Upto 10μg

A260 / A280 : 1.6 - 1.9

24) HiPurA™ Stool DNA Purification Kit (MB544)

Applications:

25) HiPurA™ Water DNA Purification Kit (MB547)

Fig. 24 a : Lane 1 to 3 : Human Stool DNA

Fig. 25 a : DNA form drainage system water

1

1

2

2

3

3Product description:

for microbial detection and quantification is highly desirable for both research and water-quality testing.

involve the binding of nucleic acids to silica spin columns.

not always balanced with the goal of obtaining high-quality DNA from complex samples.

isolate high-quality, inhibitor-free DNA from diverse water samples.

applications such as restriction enzyme digestion, PCR amplification and Southern blotting.

Principle / Technology : Spin column based protocol.

Format available : Miniprep

Sample material : 10-15 ml (or desired volume) of water sample

Elution volume :100μl

Applications:

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Mycobacterium tuberculosis is most hazardous and fatal. Multiplication time of the organism is very slow resulting in late diagnosis. This demands the need of the diagnostic tool which is able to detect very low manifestation of the infection.

however DNA is difficult to obtain from the M.tuberculosis.

M.tuberculosis has a coating on outer cell wall of mycolic acid compounds which makes it hard and resistant to many antibiotics and is a key factor for virulence.

Mycobacterium tuberculosis DNA Purification Kit isolates the DNA from M.tuberculosis with the use of heat lysis and bead beating to break open the cell wall and release DNA.

Proteinase K chews up the other protein content.



Sample material : 5ml culture suspension

Elution volume : 50μl - 100μl


A260 / A280 : 1.6 - 1.9

26) HiPurA™ Mycobacterium tuberculosis DNA Purification Kit (MB545)

1 2

Fig. 26 a : Lane 1 & 2 : Mycobacterium tuberculosis DNA

Applications:

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27) HiPurA™ Yeast Plasmid DNA Purification Kit (MB550)1 2 3

Fig. 27 a : Lane 1: 1kb ladder, Lane 2-3 : Yeast plasmid DNA


plasmids have been extensively studied in yeast anddeveloped into yeast cloning vectors. These plasmids have alsobeen used as “simple systems” to understand themechanism and control of DNA replication in eukaryotic cells.

yeast. Generally, they all involve a step to remove the yeast cell wall, followed by lysis of the cells and isolation ofplasmid DNA.

TM Yeast Plasmid DNA Purification Kit is a straightforward solution for extraction of plasmids from yeast. High efficiency enzymatic removal (using lyticase/zymolase) of the cell wall is followed by cell lysis and plasmid is purified over a spin column. The plasmid yields depend on copy number, yeast strain and growth conditions.

molecular biology procedures such as digestion withrestriction enzymes, cloning, PCR, transformation into E. coli, transfection, in vitro translation, blotting and sequencing.


Format available :Miniprep

Sample material :<2 x 107 cells

Elution volume :50μl

A260 / A280 : 1.7 - 1.8

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28) HiPurA™ Yeast Genomic DNA Purification Kit (MB552)

Fig. 28 a : Lane 1 to 3 : DNA from S. cerevisiae

1 2 3Product description:

for the DNA to be extracted. Therefore, they are converted

cell with out the cell wall yet intact without loosingcytoplasmic integrity.

zymolase or lyticase leads to spheroplasts formation.

extraction processes is carried out like complete lysis, binding to the column followed by washing and elution.

protocol; spheroplasting in log phase is easier than instationary culture.


Format available : miniprep format

Sample material : Upto 1×108 cell in log phase culture


A260 / A280 : 1.7 - 1.8

Applications:

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frozen blood, bacteria (Gram positive and Gram negative), tissues and cells are possible using HiPurA™ Multi-sample DNA Purification Kit.

different lysis solutions and specific procedure for lysis of various samples.

isolated by spin column method.

processing and ordering multiple kits.

Principle / Technology : Silica column based protocol.

Format available: Miniprep

Sample material : Blood Bacterial Animal Cultured tissue cell

Elution volume : 200μl 200μl 200μl 200μl

Total DNA yield : 4-12μg 4-20μg 30-45μg 10-20μg

A260 / A280 : 1.6 - 1.9

Applications:

Fig. 29 a : Lane 1 to 3 : DNA from goat tissueLane 4 : 1kb DNA ladder

L Fig. 29 b : Lane 1 : 1kb DNA ladderLane 2 & 3 : DNA from Chicken tissue

29) HiPurA™ Multi-Sample DNA Purification Kit (MB554)

1 2 3 4

1 2 3

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Molecular Biology

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due to polysaccharides and polyphenol compounds present within the thalli of many species.

isolation methods are often difficult to perform since these plant material release viscous soluble polysaccharides that may clog the column and pose a problem in downstream applications.

protocol, which is an easy method for DNA isolation. CTAB forms complexes with polysaccharides at salt concentrations above 0.5 M at room temperature. At lower saltconcentrations, CTAB complexes with nucleic acids.

to bind and precipitate polyphenolics. Chlorophyll and some denatured proteins are removed from the plant tissue in an organic chloroform/isoamyl step and the organic phase is separated by centrifugation

Southern blotting and analysis by PCR.

Principle / Technology : Solution based protocol.

Format available : Miniprep

Sample material :250-300mg


A260 / A280 : 1.7 - 1.8

30) HiPurA™ Algal DNA Purification Kit. (MB561)

Fig. 30 a : Lane 1 & 2 : DNA from Red algae speciesLane 4 & 5 : DNA from Brown algae speciesLane 3 : Hind III Marker

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in all cells, and is found in animals and plants, and therefore in our food.

become a useful instrument to assess the safety, quality and integrity of the food chain. This kit is designed for purification of DNA from variety of raw and processed food matrices.

allergenic material, the detection of adulteration (e.g. species replacement as a result of commercial fraud), and theidentification of microbes that cause food-borne diseases.

including researchers, food business operators and regulatory authorities. The latter may use this methodology to assess the safety and authenticity of foodstuffs, and uncover any breaches of labelling legislation.

gel, but it is PCR amplifiable and can be processed for further downstreaming operations .


Format available :Miniprep

Sample material :100-200mg of ground sample, 400 μl of liquid sample

31) HiPurA™ Food DNA Purification Kit. (MB562)

Fig. 31 a : PCR data of 16s rRNA using extracted DNA from various food sampleLane 1 : 100 bp DNA LadderLane 2 & 3 : whole milk DNA Lane 4 - 6 : DNA from milk powder Lane 7 & 8 : DNA from fruit juice Lane 9 & 10 : DNA from fruit Jam

1 2 3 4 5 6 7 58 9 10

Applications:

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Product description:TM Reagent is a ready to use reagent for

simplified isolation of RNA from human, animal, plant, yeast, bacterial and viral samples. The protocol is rapid and permitsisolation of RNA from large number of samples of small or large volumes.

phenol in a mono-phase solution, effectively dissolves RNA.

separates into 3 phases: an aqueous phase containing the RNA, the interphase containing DNA and an organic phase containing proteins.

to the single-step RNA isolation using phenol and guanidine isothiocyanate developed by Chomczynski and Sacchi.

Principle / Technology :Solution based protocol

Sample material :1ml culture or 50-100mg tissue, 5-10x106 cell

Total DNA yield : Tissue : 1-10 g depending on the type of tissueCultured cells : 3-15 g depending on the type of cells

A260 / A280 :1 .8 to 2.1

32) RNA-XPressTM Reagent (MB601)

Applications:

The purified RNA can be used in a wide range of downstream applications:

in vitro translation

1 2 3

Fig. 32 a 1 : Lane 1 to 2 : RNA from cultured cells2 : Lane 1 to 3 : RNA from plant (leaf material)

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Product Description:TM Total RNA Miniprep Purification Kit can be used

for purification of total RNA from animal cells, animal tissues, bacteria and yeast, and for cleanup of RNA from crude RNA preparations and enzymatic reactions with spin-column procedure.

and denaturation; samples are centrifuged through a HiShredder which removes insoluble material and reduces the viscosity of the lysate by disrupting viscous material.

selective binding of RNA to the silica membrane. After the initial binding of RNA, impurities like proteins, polysaccharides, low molecular weight metabolites and salts are removed by short washing steps. High quality RNA is finally eluted in the elution buffer provided with the kit.


Formats available :Miniprep

Sample material : 1 x 107 cells, 15-30mg of the animal tissue.

Elution volume : 30-50 μl

Total DNA yield. : Upto 100μg

A260 / A280 : 1.8 - 2.1

33) HiPurA™ Total RNA Miniprep Purification Kit (MB602)

Applications:

The RNA obtained is compatible with the following downstream applications

1 2 3 4

Fig. 33 a : Lane 1 to 4 : RNA from Jurkat cells

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Molecular Biology

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and filamentous fungi with spin-column procedure.

disruption and denaturation of the plant material.

separates into 3 phases: an aqueous phase containing the RNA, the interphase containing cell debris and DNA and an organic phase containing proteins.

are added, which promote selective binding of RNA to the silica membrane.

polysaccharides, low molecular weight metabolites and salts are removed by short washing steps. High quality RNA is finally eluted in the Elution Solution.



Sample material : 100mg of the plant tissue


Total DNA yield : 15-65μg

A260 / A280 : 1.8-2.1

34) HiPurA™ Plant and Fungal RNA Miniprep Purification Kit (MB603)

Applications:

The RNA obtained is compatible with the following downstream applications

Fig. 34 a : Lane 1 & 2 : RNA from A. niger culture

1 2

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by a thick cell wall which makes extraction of cellular RNA difficult. This kit simplifies isolation of yeast RNA withspin-column procedure.

helps spheroplasts formation that renders the cells vulnerable to easy lysis with detergents or rapid osmolar pressure changes.

kit helps in cell disruption and denaturation. Samples are then passed through a HiShredder which removes insoluble material and reduces the viscosity of the lysate by disrupting viscous material.

selective binding of RNA to the silica membrane.

polysaccharides, low molecular weight metabolites and salts are removed by short washing steps. High quality RNA is finally eluted in the elution solution provided with the kit.



Sample material : Upto 30 million yeast cells.



A260 / A280 : 1.8-2.1

35) HiPurA™ Yeast RNA Purification Kit (MB611)

Applications:

Fig. 35 a : Lane 1 & 2 : Yeast RNA from S.cereviseae

1 2

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bacteria (Gram positive and Gram negative) with spin column procedure.

phase is preferred as this is the only phase where theformation of the RNA and their respective translation is maximum.

elimination of the genomic DNA from the lysate. The lysate is then passed through HiShredder which removes insoluble debris material and reduces the viscosity of the lysate by disrupting viscous material.

selective binding of RNA to the silica membrane

polysaccharides, low molecular weight metabolites and salts are removed by short washing steps. Elution is done inElution Solution provided with the kit.


Formats available :Miniprep

Sample material : Upto 1x107 bacterial cells



A260 / A280 : 1.8 - 2.1

36) HiPurA™ Bacterial RNA Purification Kit (MB613)

Applications:

1 2 3 4

Fig. 36 a : Lane 1 & 2 : RNA extracted from S. aureusLane 3 & 4 : RNA from Salmonella spp.

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Product Citations

DNA Isolation Kits

Pichiapastoris. Applied Biochemistry and Biotechnology, 2013.

2. Siddarth, M., et al., Association of CYP1A1 gene polymorphism with chronic kidney disease: A case control study. Environmental Toxicology and Pharmacology, 2013. 36(1): p. 164-170.

3. Ankolkar, M., et al., Methylation status of imprinted genes DLK1-GTL2, MEST (PEG1), ZAC (PLAGL1), and LINE 1 elements in spermatozoa of normozoospermic men, unlike H19 imprinting control regions, is not associated with idiopathic recurrent spontaneous miscarriages Fertility and Sterility, 2013. 99(6): p. 1668-1673.

4. Mustafa, M., et al., Genetic polymorphisms in Cytochrome P 4501B1and susceptibility to idiopathic preterm labor in North Indian population. Clinical Biochemistry, 2013.

5. Revathy, T., M.A. Jayasri, and K. Suthindhiran, Anti-Oxidant And Enzyme-Inhibitory Potential of Marine Streptomyces. American Journal of Biochemistry and Biotechnology, 2013. 9(3): p. 282 290.

HiPurATM Quick Gel Purification Kit

1. T, P., et al., Cloning and Expresssion of Bos indicus interleukin-4 in mammalian. Indian Journal of Experimental Biology, 2013. 51: p. 352-356.

HiPurATM Plasmid DNA Miniprep Purification Kit

Asian Journal of Medical Sciences, 2009. 1(2): p. 57 63.

RNA Isolation Kits1. Gupta, S.M., et al., Cloning and characterization of GPAT gene from Lepidiumlatifolium L.:-a step towards

translational research in agri-genomics for food and fuel. Molecular Biology Reports, 2013.

2. Barui, A., et al., Assessment of Molecular Events during in vitro Re-epithelialization under Honey-Alginate Matrix Ambience. Materials Science and Engineering: C, 2013.

3. Saritha, S., K.P. Kumar, and G.R. Reddy, Reversal effect of monoisoamyldimercaptosuccinic acid (MiADMSA) for arsenic and lead induced perturbations in apoptosis and antioxidant enzymes in developing rat brain. Neuroscience, 2013.

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Electrophoresis is defined as the movement of ions and charged macromolecules through the medium when an electric current is applied. Agarose and polyacrylamide are the primary stabilizing media used in the electrophoresis of macromolecules. Agarose is mainly used for the nucleic acid electrophoresis; polyacrylamide is used for the protein electrophoresis.

Nucleic Acid Gel Electrophoresis

Agarose Gel : Agarose gel is a macroporous matrix whichallows rapid diffusion of high molecular weight (1000kDa range ) macromolecules without significant resistance. It is a non-toxic polysaccharide with high gel strength whichallows the use of concentration 1% or less. Agarose gel are

temperature permits easy recovery of samples even one which are heat labile like DNA/RNA.

For the DNA electrophoresis 0.5-1% gel of low EEO agarose (MB002) is used in submerged horizontal electrophoresis and the buffer system, usually used is TAE (ML016, ML010) or TBE.(ML017, ML022). With the change in concentration of gel and the type of buffer DNA upto 20-50000bp can be separated. For larger bp DNA, TAE buffer system is the best choice whereas TBE is preferred for the small DNA molecules (<1kb).

There is change in the buffer system used for electrophoresing RNA due to its molecular composition and the size of it being smaller than DNA. RNA is likely to form secondary structure which makes it difficult to migrate through the agarose gel matrix therefore a denaturing system is most frequently used which contains Formaldehyde, MOPS and Formamide. The MOPS buffersystem (ML050) in combination with Formaldehyde creates and a denaturing environment avoiding secondary structures of the sample loaded and its easy separation. For all the RNA works it is recommended to use DEPC treated water (ML024) for preparing buffers to inhibit the RNase action as DEPC inactivates the RNase completely.

For loading of the DNA/RNA on the gel and tracking the run, it isimportant to avoid the loss of the loaded sample and itsseparation; various DNA & RNA loading dyes serve thepurpose. Loading buffers increase the density of the sampleensuring that DNA/RNA settles evenly into the well, add colour to the sample which visually simplifies loading, they contain dye like

(ML015), Orange G (ML092)which under the electric field migrate towards anode, enablingelectrophoretic processes to be monitored.

eg. Glycerol based dye, sucrose based dye, alkaline buffered dye.

At the end of the electrophoretic run there has to bevisualization of the DNA/RNA sample separated in the processes; but for which our normal vision is not sufficient. The DNA bands

there are special fluroscent dye which associate themselves to

bromide (MB074) , which detects both single and double strand

available like Silver stain (ML123), Propidium Iodide (ML067),non hazardous SYBr Green staining (ML053) etc.

Buffers and Reagents for Electrophoresis

Fig. 37 a : Gel stained using Hi-SYBr Safe Gel Stain ML053

Fig. 37 b : DNA Gel electrophoresis using diffrent loading dyes Lane 1 : ML015

Lane 2 & 3 : ML092Lane 4 & 5 : ML086

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Fig. 37 d : Serially diluted E.coli cell extracts were loaded for SDS-PAGE and the protein bands were stained using Zinc Stain solution

Fig. 37 c : Serially diluted E.coli cell extracts were loaded for SDS-PAGE and the protein bands were stained

Protein Electrophoresis :

All analytical protein electrophoresis is carried out inpolyacrylamide gels under the condition that ensuresdissociation of proteins into their individual polypeptide subunits and that minimize aggregation .The strongly anionic detergent SDS is used in combitnation with a reducing agent and heated to dissociate proteins before they are loaded onto the gel. The denatured polypeptide binds to SDS and become negatively charged. Because the amount of SDS is almost always proportional to the molecular weight of the polypeptide and is independent of its sequence. SDS-Polypeptide complex migrates through polyacrylamide gels in accordance with the size of the polypeptide.

Polyacrylamide gel is composed of chains of polymerizedacrylamide that are cross linked by a bifunctional agent. Acrylamide/Bis-acrylamide solutions are prepared according to the rotio of thier weight (ML036-38) .

This electrophoresis is carried out discontinuous buffer system in which the buffer in the reservoirs is of a pH and ionic strength different from that of the buffer use to cast the gel. The sample passes through two gels, initially through high porosity stacking gel which has Tris-Cl (pH6.8) (ML040), and then the sample isdeposited on the thin zone on the surface of resolving gel of low porosity which contains Tris-Cl (pH8.8) (ML039). The ability of the unique buffer system to concentrate all of the SDS-Polypeptide complex in the sample into a very small volume greatly increases the resolution of gel.

Reservoir buffer contains Tris-glycine (pH8.3) (ML042) whereinteraction of Cl- ions and glycine molecules helps in migration of the sample. All the components of the system contains 0.1%SDS which is said to be Laemmli buffer system. The variation in the pH

of Tris-Cl in the stacking and resolving gels renders the porosity and ionization of the glycine molecules for better

(MB026)accelerates polymerization of acrylamide and bisacrylamide by catalyzing the formation of free radicles from Ammonium persulphate (MB003).

The protein Loading buffer is also a reducing buffer which

(ML021) is very common. To visualize the protein various type of staining reagents are available like Coomassie Brilliant blue (ML046), Zinc stain (ML057), Destaining free coomassie stain(ML091) is also availabe and these type of staining solution are used to avoid the hazardous solution of destaining reagents.

HiMedia supplies every component used and required for the above electrophoresis and its various downstream applications.

Each product is of highest purity, molecular biology grade, and stringently QC tested.

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For Molecular Biology research experiments highest quality chemicals are a prerequisite to ensure exact and reproducible results as traces of chemical impurities can interfere in the results or during the analysis. HiMedia provides a broad range of ultrapure, high quality Molecular Biology grade chemicals which can be used for all fundamental experimental methods in the field of Molecular Biology. MB grade chemicals are initially evaluated for the following aspects :

Physical parameters : All chemicals are initially tested for

specifications.

Chemical Parameters : Each and every Molecular Biology grade chemical is tested for chemical parameters at the ultra modern testing facility. They are evaluated for various molecular biology applications.

Nuclease Testing : Chemicals are tested for the presence of enzyme impurities such as DNases, RNases, and proteases as these chemicals may be used for various Molecular Biology applications.

Functionality Test : All chemicals are functionally evaluated i.e. performing a protocol with that chemical as the variable in an assay where the chemical is utilized in several laboratory applications including the isolation of DNA, RNA and proteins.

The entire range of chemicals can be divided into the following categories based upon their applications:

(i) Nucleic acid Extraction and Electrophoresis: Nucleic acid extraction is one of the most basic requirements in Molecular Biology. The diverse nucleic acid extraction protocols are of two categories:

(a) Organic extraction-based Traditional Method

(b) Commercial Column-based Method

(a) Organic extraction-based Traditional Method:

The conventional organic extraction-based alkaline lysis method and the phenol chloroform extraction strategy are very popular because they are inexpensive and do not require state-of-the-art equipments. Conventional methods consist of a lysis procedure to fragment the complex starting material (e.g., blood or tissue) and inactivate the cellular nucleases with preservation of the target nucleic acid. Usually detergents like SDS (MB010), Triton

(MB032), Guanidinium salts (MB014, MB015), and other chaotropes are used for these purpose. Reducing agents [2-Mercaptoethanol (MB041), Dithiothreitol (MB070)] prevent oxidative damage of nucleic acids. Lithium chloride (MB038) is often used specifically for extraction of RNA because Li does not precipitate double-stranded DNA, proteins or carbohydrates. Salt is essential for DNA precipitation because its cations counteract the repulsion caused by the negative charges of the phosphate backbone. Ammonium acetate (MB033) is useful because it is volatile and easily removed and at high concentration it

selectively precipitates high molecular weight molecules. Phenol solubilizes and extracts proteins and lipids to the organic phase by sequestering them away from nucleic acids. Phenol titrated to a pH of 8 (MB082) is used to separate DNA from proteins and lipids, since DNA is insoluble in basic phenol. Removal of proteins from nucleic acids can be achieved by extraction with phenol:chloroform solutions (MB078).

(b) Commercial Column-based Method:

To avoid handling of hazardous chemicals like phenol and chloroform and for making the extraction procedure simpler many companies came up with column-based kit. These kits consistently give good yield of DNA which are pure enough for downstream applications. Four simple steps are followed during the extraction procedure: lysis, binding to column, washing of column and elution. The lysis is done with a buffer containing chaotropic salts which include Guanine hydrochloride (MB014), Guanidine thiocyanate (MB015), urea (MB032) and detergents

(MB143). The chaotropic salts along with alcohol [Diluent for DNA Extraction (MB228), Isopropanol (MB063)] plays a vital role in binding of the nucleic acids to the silica-based column. Ethanol is used for 1 – 2 times to remove the chaotropic salts which is crucial to get pure DNA or RNA. In the final step, DNA/RNA is eluted from the silica membrane.

The extracted nucleic acids are electrophoresed on agarose gels (refer to the agarose section).

(ii) Protein Extraction, Purification and Electrophoresis:Efficient cell lysis and maximum protein extraction yields are vital to high-quality recombinant protein purification. Detergent-based cell lysis method has become very popular these days. In general, nonionic and zwitterionic detergents are milder, resulting in less protein denaturation upon cell lysis than ionic detergents and are used to disrupt cells when it is critical to maintain protein functions or interactions. CHAPS (MB084), a zwitterionic

commonly used for these purposes. In contrast, ionic detergents are strong solubilizing agents and tend to denature proteins, thereby destroying protein activity and function. SDS (MB010), an ionic detergent that binds to and denatures proteins, is used extensively during protein extraction and purification. During protein extraction procedure, protease inhibitors e.g. Aprotinin (MB119), PMSF (MB144) are used for the instantaneous protection of proteins from various cellular proteases.

Affinity based recombinant protein purification is emerging as purified protein is the basic requirement for proteomics. Protein A Sepharose (MB112) functions as an immunoadsorbent and is widely used for regular purification of antibodies. Glutathione reduced (MB166) and Imidazole (MB019) are used during the elution steps of GST-tagged and Nickel-based affinity chromatog-raphy, respectively.

After the extraction/purification of protein sample, it should be analyzed by electrophoresis. SDS-PAGE (SDS-Polyacrylamide

Molecular Biology Chemicals

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Gel Electrophoresis) is the most common method for analyzing protein samples through resolution on a gel according to the size. In SDS-PAGE, Acrylamide (MB068), along with Bis-acrylamide (MB005) (according to a particular ratio) forms a crosslinked polymer network in presence of Ammonium persulfate (APS, MB003). This polymerization reaction is catalyzed by TEMED (MB026). After the completion of electrophoresis, the gels are stained with coomassie dye based staining solution.

(iii) Solvents: Organic solvents are organic compounds that contain carbon atoms. These solvents have various applications which include several types of chromatography like HPLC, TLC etc., nucleic acid extraction and precipitation. 1 and 2 Butanol (MB055, MB056), Acetic acid (MB052) are used in TLC. Diluent for DNA Extraction (MB228) or Isopropanol (MB063) are used to precipitate nucleic acids in presence of salts. Chloroform (MB109), DMF (MB036), DMSO (MB058), Formaldehyde (MB059), Isoamyl alcohol (MB091), Methanol (MB154), Isopropyl alcohol (MB063) are extensively used as solvent in dissolving various chemicals required for several Molecular Biological applications. Glycerol (MB060) and DMSO (MB058) are used as cryoprotectant in freezing cell cultures.

(iv) Enzymes: Molecular Biological procedures, namely DNase I (MB085), Lysozyme (MB098), Lyticase (MB99), Proteinase K (MB096), RNase A (MB087). Lysozyme digests cell wall components of gram-positive bacteria. Proteinase K cleaves glycoproteins and inactivates (to some extent) RNase/DNase in SDS solutions. Lyticase is used in protoplast production from yeast cells.

(v) Dyes and Stains: Dyes and stains have varying applications in several Molecular Biology experiments. Dyes like Bromophenol blue (MB123) and Orange G (MB168) are basically used to track the movement of DNA or RNA during agarose gel electrophoresis. Amido Black 10B (MB165) staining solution is used for the detection of minute quantities of protein after

transfer to nitrocellulose membrane. Ethidium bromide (MB071) is an intercalating agent and used as a fluorescent tag to stain nucleic acids. It fluoresces with an orange colour under ultraviolet light and it intensifies almost 20-fold after binding to DNA. Coomassie Brilliant Blue G250 (MB092) and R250 (MB153) are used to stain proteins on polyacrylamide gels.

Stains are used in various cell biological as well as microbiological assays. DAPI (MB097) is extensively used in fluorescence microscopy as it can penetrate live as well as fixed cell membrane. Propidium iodide (MB139) is used as fluorescent stain for DNA in fixed cells during FACS analysis.

(vi) Good’s Buffer: Good s Buffers were first developed by Norman Good and colleagues in 1966 based upon several characteristics that has made them an integral part of biochemical research. This specialized group of buffer includes MES (MB020), PIPES (MB022), ACES (MB001), BES (MB118), TES (MB027), HEPES (MB016), Tricine (MB028) and Bicine (MB133) which are non toxic and has a pKa value between 6.0 and 8.0. All these buffers are highly soluble in water and have very low absorbance between 240 nm and 700 nm.

(vii) Antibiotics: Antibiotics have a wide range of applications in Molecular Biology as these are used in media while growing recombinant strains (harboring plasmids) to maintain selection pressure. The most common antibiotics are Ampicillin sodium salt (MB104) and Kanamycin sulphate (MB105).

(viii) Chromatography: Chromatography is a process for separating biomolecules which is widely used in different Molecular Biology applications. Carboxymethylcellulose (MB138) acts as a cation exchanger and DEAE Cellulose – 52 (MB110) acts as an anion exchanger. Anion exchanger has positively charged groups that attract negatively charged molecules and thus separate anionic molecules.

1A 2A 3A 4A

1B 2B 3B 4B

Fig. 38 a : All MB grade chemicals are tested for the presence of nucleases.

nuclease detection test. 4A & 4B indicate the chemical is -ve for both RNase & DNase and hence passes the nuclease detection test.

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Product CitationsAgarose

Biotechnology, 2013.

2. Koti, B.A., M. Shinde and J. Lalitha, Repeated batch production of agar-oligosaccharides from agarose by an amberlite IRA-900 immobilized agarase system. Biotechnology and Bioprocess Engineering, 2013. 18(2): p. 333-341.

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Affinity-based protein Purification

Protein purification is an essential pre-requisite for proteomics. The pure protein of interest should be free from other proteins and crude cell lysate. Protein purification procedures can vary from simple one-step precipitation procedures to large scale production processes. The most accepted of these methods isaffinity purification where the protein of interest is purified through its specific binding properties to an immobilized ligand.

Affinity tags are very efficient tools to purify heterologousproteins from different sources because the single steppurification process involves mild elution conditions and do not interfere with the structure and function of the recombinant protein. Affinity chromatography is a powerful procedure to purify IgG which is based upon the affinity of IgG to immobilizedProtein A/G agarose. In this method Protein A/G is covalentlycoupled to agarose to prepare an affinity matrix. When a suspension (e.g. serum, ascites fluid, tissue culture supernatant) containing mixture of substances along with IgG are loaded on the column (prepared from Protein A/G agarose), the IgG binds to Protein A/G and is recovered by elution.

For the purification of recombinant proteins, additional amino acids or a whole protein is often added and is known as fusion tags. One of the most common fusion tags is GlutathioneS-transferase (GST), which binds to reduced Glutathione.

Protein Crystallization

of life sciences for determining the protein structure. In 1962 the Nobel Prize in Chemistry was awarded to the pioneering studies

are diffracted by crystals.The first step during determination of

crystal which is a three-dimensional periodic arrangement of protein molecules. Protein crystallization is the most commonly used technique for resolving protein structures at the atomic level which in turn enables to understand the protein function. This entire study helps in designing novel drugs that target a particular protein.

While crystallizing a protein sample, it should be made sure to be as homogenous as possible with a purity of greater than 95%. Several conditions come into factor if a protein sample will crystallize or not. Some of these factors include protein purity and concentration, pH, temperature, precipitants and additives. pH conditions are very important due to the fact that different pH can result in different packing orientations. Precipitants, such as ammonium sulfate or polyethylene glycol (PEG) are compounds that cause the protein to precipitate out of solution.

Fig 39 : Purified IgG (using Protein G agarose) on SDS-PAGE

Lane 1: Protein Molecular Weight Marker

Lane 2: Purified IgG

Lane 1

97.4kD

66kD

43kD

29kD

20.1kD

14.3kD

Lane 2

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HiMedia introduces Nucleo-Proteo cards, a remarkably easy way to collect and isolate nucleic acid samples for analysis.

matrix, and the nucleic acids are instantly captured and stabilized.

analyze whenever you are ready.

Hi705™ Cards

the DNA is released and unravels into the matrix

Strengths:

Hi-ProPRIME™ Cards

cells lyse. As cell is lysing, the DNA is released and unravels into the matrix.

Strengths:

acids for long term ambient applications

REQUIRED

Nucleo-Proteo cards

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Agglutination is a reaction of clumping together of antigen-bearing cells, microorganisms or particles in the presence of specific antibodies (agglutinins) in a suspension. Reaction time for agglutination to occur is shorter compared to other antigen-antibody interactions. Latex agglutination makes use of latex particles which are built from different organic materials to a desired diameter, and may be functionalized with chemical groups to facilitate attachment of molecules. Latex agglutination test is based on an agglutination reaction between antigen and antibody for the detection of a pathogen. This is a very popular test in clinical laboratories. These tests have been applied to the detection of several infectious diseases. The first instance of a test based on latex agglutination was the Rheumatoid Factor Test proposed by Singer and Plotz in 1956. Since then, tests to detect microbial and viral infections, autoimmune diseases, hormones, drugs and serum proteins have been developed and marketed worldwide. It can be used for detection of both antigen and antibody.

The principle of latex agglutination is similar to that of haemagglutination but it is more sensitive due to the use of smaller, antibody-coated latex particles. These tests are designed for fast, economical, simple and accurate identification of variety of diseases. Furthermore, these are extremely simple and inexpensive tests which can be done within the capability of most laboratories.

This pathogen detection method requires no expensive orsophisticated equipment and specific expertise. These methods employ latex particles, which efficiently adsorb proteins at alkaline pH. Specific antibodies against bacterial antigens are coated on the latex particles and are then used for slide agglutination. Interaction between specific antibodies with the antigen results in visible agglutination in contrast to other suspensions that remains unagglutinated. The antibodies are adsorbed to the latex particles by binding to the Fc region of antibodies leaving Fab region free to interact with antigen present in the applied specimen. The use of smaller latex particle has improved the sensitivity and reagent longevity of latex agglutination.

Although many advanced immuno-techniques are now available, latex agglutination remains advantageous for the following reasons:

1. It requires no special equipment or trained personnel for its performance.

2. It is more sensitive than methods like counter immunoelectrophoresis for detection of soluble polysaccharide antigens in body fluids.

3. A large number of samples can be tested and the results are accepted to be confirmatory.

4. They have overcome the inherent overnight delay associated with the culture method for identification of bacteria.

5. Can be carried out as POC (Point of Care) testing, e.g., by the

6. It is approved by the World Health Organization (WHO) for detection of pathogens.

In an effort to encompass the new exciting dimensions in the field of diagnostic methods, these kits are introduced for rapid serogrouping of several clinically relevant pathogenic bacteria, thus assisting faster patient management.

Applications of Latex agglutination test kits:

Listeria, Rotavirus etc).

Storage and Shelf life:

Latex agglutination test kit can be stored for 13 months when stored at 2-8oC.

Latex Agglutination Test Kits

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An abnormal blood count or blood cell morphology does not necessarily indicate a primary hematology problem, as they may reflect an underlying non-hematological condition or may be the result of therapeutic interventions. Anemia is common in many conditions but a primary blood disease should be considered when a patient has splenomegaly, lymphoadenopathy, a bleeding tendency, thrombosis and/or non-specific symptoms of malaise, sweats or weight loss.

Although the range of hematological tests available to support clinical and public health services is broad, it is often the simplest investigations which are most useful in indicating the diagnosis. Hence, HiMedia has developed a range of hematology kits for the screening and confirmation of sickle cell anemia and detection of Thalassemia. These are divided based upon their functionality as follows:

Sickle Cell Anemia detection:

1. Primary Screening: These kits are developed forscreening of sickle cell anemia, based on solubility difference between HbS and HbA. This range includes Hi-Super Speed Sickle Kits (MBP004, MBP006 and MBP007).

Hematology Kits2. Secondary Screening: These kits are based uponcentrifugation method which detects the differential solubility of HbS and HbA. These kits have an added advantage for partial distinction between HbS (Homozygous) and HbAS (Heterozygous) in sickle cell anemia patients. This range includes Hi-Speed Sickle Kit (MBP002 and MBP003).

3. Confirmatory Kit: This kit is intended for discrimination of hemoglobin variants using alkaline gel electrophoresis as the abnormal hemoglobins have a sufficiently altered chargedistribution and can therefore be easily identified byelectrophoresis. It is a confirmatory tool for detection of sickle cell anemia. This range includes Alkaline Hemoglobin Electrophoresis Kit (MBP001).

Thalassemia detection Kit:

1. Beta-Thalassemia Detection Kit (PCR Based)


Hematology kits have a shelf-life of one year when stored at 15-25oC and Beta-Thalassemia Detection Kit has a shelf-life of 6 months when stored at -20oC.

N – Normal

SCT – Sickle Cell Trait

SCD – Sickle Cell Disease

Fig. 40 : Hemoglobin variants separated on 0.8% Agarose gel, special Low EEO using Alkaline Hemoglobin Electrophoresis Kit

Foetal hemoglobinSCDSCTN

Carbonic anhydrase

HbA2HbS

HbA

HbS

HbF

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Ladders / Markers

DNA ladders are a mixture of specially designed double-stranded DNA fragments for determining the exact size of PCR products and engineered DNA fragments. These ladders are ready-to-use and are suitable for use as molecular weight standards for agarose gel electrophoresis. Referal bands with increased intensity are present in each ladder. Recommended load : 5 l/lane

DNA Ladders

Protein Ladders

Prestained Protein Ladder is a three-color protein standard with 12 pre-stained proteins covering a wide range molecular weights from 10 to 245 kDa when separated on SDS-PAGE (Tris-glycine buffer). Prestained Protein Ladder is designed for monitoring protein

nitrocellulose) and for approximating the size of proteins. The ladder is supplied in gel loading buffer and is ready to use. Do not heat, dilute or add reducing agent before loading. Recommended load : 5 l/lane

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Products for PCR and RT-PCR

ENZYMES REAGENTS MASTER MIX KITS

Taq Polymerase dNTPs 2X PCR TaqMixture Hi-PCR KitConcentration: 5 Units/μl

Concentration: 3 Units/μl 10 mM dNTP Mix Hi-SYBr Master Mix Hi-cDNA Synthesis KitConcentration: 1 Unit/μl

100 mM dNTP Mix Hi-Chrom PCR PCR Kits for pathogen Hi-Proof DNA Polymerase Master Mix detection (Real time &)

10 mM dNTP Set Semi Quantitative)Hi-Long Amp DNA Polymerase

100 mM dNTP Set Hi-Script one StepHi-Temp DNA Polymerase RT-PCR Kit

100 mM dATPChrom Taq DNA Polymerase

100 mM dCTPM-MuLV Reverse Transcriptase

100 mM dGTPAMV Reverse Transcriptase

100 mM dTTP T4 DNA Ligase

PCR Enhancers

MB Grade Water for RT-PCR (ML065)

Nucleic acid Amplification

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DNA PolymerasesTaq Polymerase

Description:

Taq DNA Polymerase is a thermostable DNA polymerase of thermophillic bacterium Thermus aquaticus. The enzyme catalyzes

activity. In addition, Taq DNA Polymerase exhibits deoxynucleotidyl transferase activity, resulting in addition of extra

Features:

Thermus aquaticus

at 95°C

suitable for TA cloning

deoxygenin,fluroscently-labelled nucleotides)

2SO4 [(NH4)2SO4 allows for PCR at wide range of Mg concentrations and decrease inunspecific priming]

E.coli cells with a pol gene from Thermus aquaticus

Guidelines for PCR optimization using HiMedia’s Taq Polymerase:

DNA Template

1. Use high quality, purified DNA templates

2. Approximately 104 copies are required to detect the amplification in 25-30 PCR cycles

3. Use higher DNA concentration when few PCR cycles are desired

Primers

1. Generally 20-30 bp in size

2. GC content between 40-60% ideally

3. Melting temperatures should be between 42-65°C

4. Final concentration to be used 0.1-0.5μM of each primer

Magnesium Concentration

1. Ideal for Taq Polymerase is 1.5-2.0mM

2. Optimum concentration depends on template, buffer and dNTPs

3. Higher than optimal concentration yields undesired products and if concentration is too low the concentration, no amplification products are detected

dNTPs

1. Typical concentration to be used is 200μM

2. Higher than optimal concentration of dNTPs yields higher yield but low fidelity

PCR grade water

1. It is advisable to use PCR or MB grade water for any kind of PCR assay . As this water is free of nucleases and free of nucleic acid contamination that may cause false-positive signals in PCR

Taq Polymerase

1. Typical concentration to be used is 0.5 to 2 units per 50μl of reaction

PCR reaction

1. Thaw all reaction components on ice

2. To PCR reaction, add Taq Polymerase at the end

3. Once the reaction is set, immediately transfer the tubes to pre-heated thermal cycler

4. Start the reaction with desired cycling conditions with annealing temperature set to 5°C difference of melting temperature between forward and reverse primers

Representative Data 1 2 3 4 5 6 7 8

Lane Amplicon Size Taq DNA dNTP Formamide

Polymerase Concentration Concentration

Concentration

1 M1-1Kb Ladder

2 1.5Kb 1.0U 0.2mM NA

3 5.0Kb 1.25U 0.25mM 3%

4 8.0Kb 1.25U 0.25mM 3%

5 10.0Kb 1.25U 0.36mM 3%

6 15.0Kb 1.25U 0.36mM 3%

7 20.0Kb 1.25U 0.36mM 3%

8 M2-Lambda Hind III marker

Fig. No 41 a : Figure representing amplicon sizes obtained using Taq DNA Polymerase (MBT060)

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Different types of Taq Polymerase available:

Sr.No Product Code Description Concentration

50mM MgCl2 Enzyme vial

2 5 Units / μl



2 3 Units / μl



2 1 Units / μl


Hi-Temp DNA Polymerase

Description:

Hi-Temp DNA Polymerase is a complex of specific anti-Taq monoclonal antibody with best quality thermostableTaq DNA Polymerase for automatic “hot start” amplification, resulting in greatly enhanced amplification specificity, sensitivity and yield. Hi-Temp DNA Polymerase catalyses the polymerization of

of Mg

Applications:

up to 3 kb

Hi-Proof DNA Polymerase

Description:

Hi-Proof DNA Polymerase is an extremely thermostable polymerase from the hyperthermophilicarchaeumPyrococcusfuriosus. Hi-Proof DNA Polymerase catalyzes the

direction in the presence of Mgexonuclease (Proofreading) activity that enables the polymerase to correct nucleotide incorporation errors resulting in over 10-fold higher fidelity than possible with Taq DNA Polymerases. It has no

Applications:

1 2 3 4 5 6

Lane Amplicon Size Hi-Proof DNA dNTP Formamide

Polymerase Concentration Concentration

Concentration

1 M1-1Kb Ladder

2 0.5Kb 1.25U 0.2mM NA

3 1.5Kb 1.25U 0.2mM NA

4 5.0Kb 1.25U 0.25mM 3%

5 8.0Kb 1.25U 0.25mM 3%

6 M2-1Kb Ladder

Fig. No 41 b : Figure representing amplicon sizes obtained using Hi-Proof DNA Polymerase (MBT068)

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Hi- Long Amp DNA Polymerase

Description:

Hi-Long Amp DNA Polymerase is a modified and optimized thermostable enzyme blend containing Taq DNA polymerase, Hi-Long Amp DNA polymerase and enhancing factors for high throughput PCR applications. It ensures higher sensitivity, longer PCR products and higher yields compared to conventional Taq

resulting in considerably higher amplification fidelity than possible with unmodified Taq DNA polymerase.

Applications:

Representative Data

ChromTaq DNA Polymerase

Description:

Taq DNA Polymerase is a thermostable DNA polymerase of thermophillic bacterium Thermus aquaticus. It is suitable for applications requiring high temperature synthesis of DNA. The

exonuclease activity. In addition, Taq DNA Polymerase exhibits deoxynucleotidyl transferase activity, resulting in addition of extra

Features:

of polymerase to the reaction.

ing and serves as tracking dyes during gel electrophoresis. The blue and pink color dyes migrate approximately at 4 kb and 0.3 kb respectively on 1% TAE agarose gel.

Thermus aquaticus.

40 minutes at 95°C.

suitable for TA cloning.

deoxygenin,fluroscently-labelled nucleotides).

2SO4. [(NH4)2SO4 allows for PCR at wide range of Mg concentrations and decrease unspecific priming].

Product Citations

Taq Polymerase

genes from seafood. Journal of Microbiological Methods, 2013. 93(3): p. 233-238.

2. Iyer, S.N., et al., Determination of common genetic variants in cytidinedeaminase (CDA) gene in Indian ethnic population. Gene, 2013. 524(1): p. 35-39.

3. Rauta, P.R., M. Dhupal, and B. Nayak, Screening and characterization of potential probiotic lactic acid bacteria isolated from vegetable waste and fish intestine. International Journal of Current Microbiology and Applied Sciences, 2013. 2(8): p. 234- 244.

Fig. 41 c : Figure representing amplicon sizes obtained using Hi-Long Amp DNA Polymerase with HiBuffer A and HiBuffer S.

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Properties Applications

Sr.No Name of Polymerase Product

Code

Template

Length

Hot-

Start

Proof-

reading

Blunt End

Cloning

TA Cloning High

Fidelity

PCR

Direct Gel

Loading

1Taq DNA Polymerase(Concentration: 5 U / μl)

MBT060,MBT060A

Upto 8 kb — — — + — —


MBT060B,MBT060C

Upto 8 kb — — — + — —


MBT060D,MBT060E

Upto 8 kb — — — + — —

4Hi-Proof DNA Polymerase(Concentration: 5 U / μl)

MBT068 Upto 8 kb — + + — + —

5Hi-Long Amp DNA Polymerase(Concentration: 5 U / μl)

MBT069 Upto 20 kb — — + — — —

6Hi-Temp DNA Polymerase(Concentration: 5 U / μl)

MBT070 Upto 8 kb + — — — — —

7ChromTaq DNAPolymerase(Concentration: 1 U / μl)

MBT088 Upto 20 kb — — — + — +

+ : Applicable — : Not Applicable

Selection Guide for DNA Polymerases

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Reagents and Master MixesHi-cDNA Synthesis Kit

Description:

Hi-cDNA Synthesis Kit is designed for reverse transcription where cDNA (complementary DNA) is synthesized in vitro from an mRNA template by an enzyme that has reverse transcriptase activity.

RT) is an RNA-dependent DNA polymerase that can be used in cDNA synthesis.This step is very important in order to perform PCR since DNA polymerase can act only on DNA templates. The resulting cDNA is single-stranded and this process is called reverse transcription (RT) or first strand cDNA synthesis.

Representative Data:

M-MuLV Reverse TranscriptaseDescription:

Reverse Transcriptase) is an RNA-dependent DNA polymerase requiring a DNA primer and an RNA template to synthesize a

RNaseH activity enhances the synthesis of long cDNAs and therefore the enzyme is recommended for preparing long cDNAs.

Applications:

1. First strand cDNA synthesis

2. DNA Labelling

3. RNA analysis by primer extension

Restriction EnzymesRestriction enzymes are enzymes isolated from bacteria that recognize specific sequences in DNA and then cut the DNA to produce fragments, called restriction fragments. Restriction enzymes play a very important role in the construction of recombinant DNA molecules, as is done in gene cloning experiments. Another application of restriction enzymes is to map the locations of restriction sites in DNA.

Each restriction enzyme recognizes a short, specific sequence of nucleotide bases (the four basic chemical subunits of the linear double-stranded DNA molecule—adenine, cytosine, thymine, and guanine). These regions are called recognition sequences and are randomly distributed throughout the DNA. Different bacterial species make restriction enzymes that recognize different nucleotide sequences as a part of their defense mechanism against invading viruses.

When a restriction endonuclease recognizes a sequence, it snips through the DNA molecule by catalyzing the hydrolysis (splitting of a chemical bond by addition of a water molecule) of the bond between adjacent nucleotides. Bacteria prevent their own DNA from being degraded in this manner by disguising their recognition sequences. Enzymes called methylases add methyl groups (—CH3) to adenine or cytosine bases within the recognition sequence, which is thus modified and protected from the endonuclease. The restriction enzyme and its corresponding methylase constitute the restriction modification system of a bacterial species.

HiMedia provides a range of Restriction enzymes like EcoR I, BamH I, Hind III, Hinc II, Pst I, Kpn I, Taq I and so on the list

Polymerases and Amplification

Fig. 42 b : Lane 1: 1kb DNA Ladder Lane 2 : RT-PCR Product (Test)

Fig. 42 a : Lane 1: Hind III DNA ladder Lane 2 - 4 : DNA digested using EcoR I Lane 5 - 7 : DNA digested using Hind III

1 2 3

1 2 3 4 5 6 7

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AMV Reverse TranscriptaseDescription:

Transcriptase) is an RNA-dependent DNA polymerase with a molecular weight of 157,000 daltons. This enzyme can synthesize a complementary DNA strand initiating from a primer using either RNA (cDNA synthesis) or single-stranded DNA as template.

Features:

and RNase H activities

temperatures.

Capable of synthesizing cDNA over a wide range of temperatures.

Applications:

T4 DNA LigaseDescription:

T4 DNA Ligase catalyses the formation of a phosphodiester bond

duplex DNA or RNA. This enzyme will join blunt end and cohesive end termini as well as repair single stranded nicks in duplex DNA, RNA or DNA-RNA hybrids.

Features:

hybrids

Hi-PCR KitHi-PCR Kit is designed to perform PCR on various DNA templates for numerous downstream applications. Hi-PCR kit contains

Taq Polymerase, dNTP mixture and MgCl2 for reproducible PCR results. The kit also includes two buffers for standard reactions one with and another without MgCl2 included in the buffer. The user needs to only add the desired pure template and primers of interest to perform the desired PCR.

Features:

suitable for TA cloning

deoxygenin,fluroscently-labelled nucleotides)

Hi-Chrom PCR Master MixDescription:

Hi-Chrom PCR Master Mix is a premixed ready-to-use solution containing Taq DNA polymerase, dNTPs, MgCl2 and reaction buffers at optimal concentrations for efficient amplification of DNA templates by PCR. Hi-Chrom PCR Master Mix is recommended for any amplification reaction that will be visualized by agarose gel electrophoresis and ethidium bromide staining. The master mix is not recommended if any downstream applications use absorbance or fluorescence excitation.

This pre-mixed formulation saves time and reduces contamination and pipetting volume error due to a reduced number of pipetting steps required for PCR set up. The mix is optimized for efficient and reproducible PCR.

Note: Hi-Chrom PCR Master Mix is provided as a pre-mix with gel loading dye which doesnot hinder the amplification reaction as is does not contain any inhibitors.

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Hi-SYBr Master MixDescription:

use mix convenient for real time PCR. The master mix contains:

SYBr Green Dye, Hi-Temp DNA polymerase, dNTPs, Assay buffer, MgCl2Template, primers and nuclease-free water has to be added before setting up the PCR reaction.

Due to the ready-made mixture, the reaction set time is reduced to half.

Features:

mizes non-specific amplification and primer dimer formation

errors and contamination od reaction mix

Applications:

2X PCR Taq Mixture

Description:

containing Taq DNA polymerase, dNTPs, MgCl2 and reaction buffers at optimal concentrations for efficient amplification of DNA templates by PCR. This pre-mixed formulation saves time and reduces contamination due to a reduced number of pipetting steps required for PCR set up. The mix is optimized for efficient and reproducible PCR.

Features:

Application:

Fig. 42 c : Genomic DNA (50ng, 10ng, 1ng) was amplified for 35 cycles with primers human GAPDH gene and

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PCR Based Diagnostic KitsPCR has become one of the most important tools in molecular diagnostics, providing maximum sensitivity and specificity for detection of nucleic acid targets. Real-time monitoring of PCR has simplified and accelerated PCR laboratory procedures and has increased information obtained from specimens including routine quantification and differentiation of amplification products. PCR based methods have emerged as the most sensitive and reliable molecular technique in clinical applications for pathogen detection and simultaneous identification of multiple targets. PCR based diagnostic kits facilitate rapid detection of pathogenic organisms including viruses and bacteria in clinical samples as compared to traditional antibody-based serological methods. The sensitivity of this technique allows pathologists to diagnose any infectious agent, such as Mycoplasma, E.coli, Mycobacteria, Dengue etc. at an initial stage of infection by detecting fewer copy numbers of their genetic material. Modified PCR protocols and methodologies are today available which allow quick detection and quantification of viral RNA in clinical samples. All these protocols are based widely upon the RT-PCR technology, i-e, PCR preceded by a reverse transcription in a one-step reaction. The entire range of diagnostic kits can be divided into the following categories:

1. PCR based kits (Semi-quantitative): The principle of all the kits are based upon detection of a pathogen by using semi-quantitative PCR methods. This range includes detection of E.coli O157:H7 (MBPCR002), Salmonella (MBPCR004), Mycobacteria (MBPCR009),Dengue (MBPCR011) ,etc.

2. PCR based kits (Quantitative): All these kits utilize the principle for detection of a microorganism using quantitative PCR methods. This range includes Mycoplasma (MBPCR015) and Mycobacterium (MBPCR017).


PCR based diagnostic kits have a shelf-life of six months when stored at -20oC.

MBPCR002 - E.coli O157: H7 Detection Kit

1 2 3 4 5 6 7 8 9 10 11 12 13 14 Lane Sample Lane Sample

1 - 8 E.coli O157:H7 (From Juice)

2 -ve control 9 Enterococci

3 -ve control 10 E.coli O157:H7 (From Meat)

4 E.coli O157:H7 11 P.aeroginosa

5 S.aureus 12 K.pneumoniae

6 S.faecalis 13 E.coli

7 L.monocytogenes 14 100 bp DNA Ladder

Specifications

Sensitivity: Detectable upto 10-100 CFU / ml (mg) beforepre-enrichment.

Specificity: 100% exclusivity for about 100 non-target strains.

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MBPCR003 - Vibrio cholerae Detection Kit

1 2 3 4 65

1 2 3 4 5

Lane Sample

1

2

3 Hosp. lsolate 1

4 Hosp. Isolate 2

5 -ve control

6 100bp DNA Ladder (MBT049)

Specifications



MBPCR004 - Salmonella Detection Kit

MBPCR005 - Listeria monocytogenes Detection Kit

1 2 3 4 5 6 7 8 Lane Sample

1 Salmonella pure culture

2 Salmonella (Clinical isolate)

3 Salmonella (From Juice)

4 Salmonella (From raw meat)

5 -

6 Salmonella (From raw fish)

7 -ve control

8 100bp DNA Ladder (MBT049)

Specifications:



Lane Sample

1 -ve control

2 Listeria (Pure Culture)

3 Listeria (Hosp. Isoloate 1)

4 Listeria (Hosp. Isoloate 2)

5 100 bp DNA Ladder (MBT049)

Specifications:



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MBPCR009 - Mycobacteria tuberculosis Detection Kit

1 2 3 4 5 6 7

1 2 3 4 5 6 7 8

Lane Sample

1 100 bp DNA Ladder

2 M.tuberculosis

3 Clinical isolate 1

4 Clinical isolate 2

5 M.tuberculosis control

6 -ve control

7 -ve control

Specifications:

Sensitivity: Detectable upto 10-100 CFU / ml (mg)


MBPCR008 - Methicillin Resistant Staphylococcus aureus (MRSA) Detection Kit (Uniplex)Lane Sample

1 100 bp DNA Ladder

2 -ve control

3 -ve control

4 MRSA DNA

5 Hospital Isolate 1




Specifications:


Specificity: 100% exclusivity for about 100 non-target strains

MBPCR019 - Staphylococcus Cassette Chromosome (SCC) mec typing MRSA Detection Kit (Multiplex)

1 2 3 4 5 6 7 8 9 10 11typing MRSA Detection Kit (Multiplex) is a qualitative conventional PCR kit which focuses on simultaneous amplification of

determinant as an internal control using specific primers. The amplified target is confirmed by using agarose gel electrophoresis.

Lane 1 & 11 2 3 4 5 6 7 8 9 10

Ladder SCCmec type Negative

Specifications:

Sensitivity: Detectable upto 103 CFU/ml.


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MBPCR020 - Methicillin Resistant Staphylococcus aureus (MRSA) Detection Kit (Multiplex)

MBPCR022 - Multi-Drug Resistant Mycobacterium tuberculosis Detection Kit (Multiplex)

Detection Kit (Multiplex) is a qualitative conventional PCR kit which focuses on simultaneous amplification of four targets, Staphylococcus genus (16S rRNA gene), methicillin-resistant

toxin (lukS gene), and discrimination between S. aureus and CoNS (femA gene). The amplified target is confirmed by using agarose gel electrophoresis.

Lane Sample

1 100 bp DNA Ladder

5 MSSA

Specifications:

Sensitivity: Detectable upto 103 CFU/mL

Specificity: 100% exclusivity for about 100 non-target strains

1 2 3 4 5 6 7 8 9 10 11 12

273 bp180 bp

1 2 3 4 5 6

597 bp450 bp297 bp

151 bp

Detection Kit is designed to detect mutations in rpoB (180 bp) and katG (273 bp) gene from sputum specimens of MDR-TB by PCR. Rapid diagnosis of MDR-TB plays a vital role in guiding standardized treatment regimen. Traditional drug susceptibility test (DST) takes 6-8 weeks. Even with the aid of liquid cultures and radiometry measures, the bacteriological cultures and DST still takes 2-3 weeks. Hence, to identify MDR-TB with the advance techniques like PCR becomes critical.

Lane Sample

1 50 bp DNA Ladder

12 Non Template control

Lane No. 4, 5, 8, 9 - contains no sample loaded

Specifications:

Sensitivity: Detectable upto 10-100 CFU / ml (mg)


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Lymphocyte and Granulocyte Separation media

The principle of Lymphocyte and Granulocyte Separation Media (LSM GSM) is based on the adapted method of isolating lympho-cytes or granulocytes using centrifugation techniques by Bøyum. In this specialized centrifugation technique defibrinated blood is layered on a solution of polysucrose and sodium diatrizoate and centrifuged at low speed for 30 minutes. Differential migration following centrifugation results in the formation of several cell layers. The PBMCs (Peripheral Blood Mononuclear Cells) are found in the buffy coat which is a ring-like, typically white layer of nucleated cells between the blood plasma and Lymphocyte and Granulocyte Separation Media.

Applications of Lymphocyte and Granulocyte Media:

in vitro

Isolation of lower density mononuclear cells

For studying cell mediated lympholysis and for human lymphocyte antigen (HLA) typing

For T and B-cell enumeration


Lymphocyte and Granulocyte Separation Media are stable for 3 years when stored at 2-8oC. As the medium is light sensitive it should be protected from direct light. Media are supplied in plastic bottles with screw caps. The caps are wrapped with plastic tapes to prevent any leakage.

Intended useTM LSM 1077 is an iso-osmotic, low viscosity

medium containing polysucrose and sodium diatrizoate adjusted to a density of 1.0770 ± 0.0010 g/ml. This medium offers a quick and reliable method for the simple isolation of human peripheral blood mononuclear cells (PBMC) and lymphocytes from defibrinated EDTA or heparin treated human blood. It is certified for in vitro

The method is applicable for studying cell-mediated lympholysis and for human lymphocyte antigen (HLA) typing. It may also be employed as the initial step prior to enumeration of T-, B- and

“null” lymphocytes.

Summary and principle of the procedure

The first step in studying lymphocytes is to isolate them so that their behavior can be analyzed in vitro. Lymphocytes are present in blood, peritoneal exudates or lymphoid organs mixed with other cells. Human lymphocytes can be isolated most readily from peripheral blood. A pure population of lymphocytes can be obtained by various separation procedures.

HiSep™ LSM is based on the adapted method of isolating lymphocytes using centrifugation techniques by Boyum in which diluted defibrinated blood is layered on a solution of sodium diatrizoate and polysucrose and centrifuged at low speeds for 30 minutes.

Differential migration following centrifugation results in the formation of several cell layers. Mononuclear cells (lymphocytes and monocytes) and platelets are contained in the banded plasma-LSM interphase due to their density, and the pellet that is formed contains mostly erythrocytes and granulocytes, which have migrated through the gradient to the bottom of the tube. Lymphocytes are recovered by aspirating the plasma layer and then removing the cells. Excess platelets, HiSep™ LSM, and plasma can then be removed by cell washing with isotonic phosphate buffered saline.

TM LSM 1077

Density Gradient Separation Media

Product Name Density Intended use

HiSepTM LSM 1077 1.0770 g/ml Recovery of viable human PBMC

HiSepTM LSM 1073 1.073 g/ml Isolation of lower density human mononuclear cells

HiSepTM LSM 1084 1.084 g/ml Isolation of viable mononuclear cells from chicken, rats, mice

GranuloSepTM GSM 1119 1.119 g/ml Separation of mononuclear cells and granulocytes

Ficoll 400-20% -- Commonly used to prepare density gradients

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Product Citations

GranulosepTM GSM 1119

1.Kundu, S., et al., Oxidative stress as a potential biomarker for determining disease activity in patients with Rheuma-

toid Arthritis. Free Radical Research, 2012. 46(12): p. 1482-1489.

HiSepTM LSM 1073

in lymphocytes of patients with rheumatoid arthritis.Inflammopharmacology, 2013.

2.Kundu, S., et al., Oxidative stress as a potential biomarker for determining disease activity in patients with Rheuma-

toid Arthritis. Free Radical Research, 2012. 46(12): p. 1482-1489.

3.Mukhopadhyay, D., et al., Miltefosine Effectively Modulates the Cytokine Milieu in Indian Post Kala-Azar Dermal

Leishmaniasis. The Journal of Infectious Diseases, 2011. 204.

HiSepTM LSM 1077, HiSepTM LSM 1084

1.Bala, A., et al., Carbon tetrachloride: A hepatotoxin causes oxidative stress in murine peritoneal macrophage and

peripheral blood lymphocyte cells.Immunopharmacology and Immunotoxicology, 2012. 34(1): p. 157-162.

2.Gupta, S. and C. Haldar, Physiological crosstalk between melatonin and glucocorticoid receptor modulates T-cell

mediated immune responses in a wild tropical rodent, Funambuluspennanti. The Journal of Steroid Biochemistry and

Molecular Biology, 2013.

134: p. 23-36.

3.Tyagi, A.M., et al., Estrogen Deficiency Induces the Differentiation of IL-17 Secreting Th17 Cells: A New Candidate in

the Pathogenesis of Osteoporosis Plos One, 2012. 7(9).

Mice: A Key Mechanism for Anti-Osteoclastogenic Effect Plos One, 2011. 6(6).

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Horizontal Electrophoresis Apparatus and Power Packs Hi-Eco Mini Horizontal Gel Box LA844Features

Robust acrylic frame mounted on non-slip rubber feet.

A safety lid fits on one way only preventing access to live components.

Fully shrouded connectors on safety lid.

Strong thick removable gel casting tray resists warping.

End gaskets allow gel pouring in the running tank or in an external casting unit.

Long life pure platinum electrodes.Convenient carrying handles enable easy buffer disposal and increased mechanical strength.

Operating temperature range 4-65°C.

Sr No Name of Item Specifications

1. Principal Material Acrylic2. Inner Tank Dimension L x W x H 16.2 x 9.9 x 2.9 cm3. No. of combs 2 Nos.4. No. of gel casting tray 1 no.5. Connecting cord 1set (black and 1Red)6. No. of platinum electrodes 1 anode 1 cathode7. Lid 1 No.8. Recommended power supply LA842

10. Comb thickness 1.5mm polycarbonate11. Comb configurations 8 tooth, 8 tooth

13. Max. Buffer volume 150 ml

Ordering Information

Hi-Eco Mini Horizontal Gel Box LA844

Mini Horizontal LA665Electrophoresis SystemFeatures






Long life pure platinum electrodes.

Convenient carrying handles enable easy buffer disposal and increased mechanical strength.



1. Principal Material Acrylic2. Inner Tank Dimension L x W x H 17.6 x 12 x 9.5cm3. No of combs 2 Nos. 4. No. of gel casting tray 1 no.5. Connecting cord 1set (black and 1Red)6. No. of platinum electrodes 1 anode, 1 cathode7. Lid 1 No.8. Recommended power supply LA6909. Gel Dimensions 7 x 8 cm (gasketed)10. Comb thickness 1.5mm polycarbonate11. Comb configurations 6 & 10 tooth



Mini Horizontal LA665Electrophoresis System

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Horizontal Gel Box LA884Features










1. Principal Material Acrylic2. Inner Tank Dimension L x W x H 24.5 x 18 x 9.5 cm3. No of combs 2 Nos.4. No. of gel casting tray 1 no. 5. Connecting cord 1set (black and 1Red)6. No. of platinum electrodes 1 anode 1 cathode7. Lid 1 No.8. Recommended power supply LA6909. Gel Dimensions 12x14cm (gasketed)10. Comb thickness 1.5mm polycarbonate11. Comb configurations 12 & 20 tooth



Mini Horizontal Gel Box LA884

Horizontal Electrophoresis System LA666Features










1. Principal Material Acrylic2. Inner Tank Dimension L x W x H 22 x 15 x 9.5cm3. No of combs 2 Nos.4. No. of gel casting tray 1 no.5. Connecting cord 1set (black and 1Red)6. No. of platinum electrodes 1 anode, 1 cathode7. Lid 1 No.8. Recommended power supply LA6909. Gel Dimensions 9 x 11cm (gasketed)10. Comb thickness 1.5mm polycarbonate11. Comb configurations 10 & 14 tooth

13. Max. Buffer volume` 600 ml


Horizontal LA666Electrophoresis System

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100 well Multi channel compatible Horizontal Gel Box LA839Features










1. Principal Material Acrylic2. Inner Tank Dimension L x W x H 36 x 29.5 x 8cm3. No of combs 4 Nos.4. No. of gel casting tray 1 no.5. Connecting cord 1set (black and 1Red)6. No. of platinum electrodes 1anode 1 cathode7. Lid 1 No.8. Recommended power supply LA6909. Gel Dimensions 25 x20 cm10. Comb thickness 1.5mm polycarbonate



100 well Multi channel LA839 compatible Horizontal Gel Box

Electrophoresis Power Supply LA842Features

Two apparatus can be connected to the power supply with dual output at both side of the power supply.

After switching ON the mains supply power, red led will glow.

front panel.


Selectable, 200 mA

3. Maximum oprating temprature 45°C4. No. of output 2 nos5. Dimension in mm L x W x H 105 x 125 x 656. Recommended for LA844


Electrophoresis Power Supply LA842(2 Terminal out put)

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Electrophoresis power supply LA690Features

Safety features including automatic detection and auto shut-off in case of 1) no load 2) ground leak and short circuit 3) cooling fan failure 4) system overheating.

Extremely silent operation.

Highly cost-effective model.

No need of voltage stabilizer as the unit works on the advanced technology of Switch Mode Power Supply (SMPS).


2) Constant current 0 to 400 mA3. LED digital display of parameters4. 4 output terminals that can run upto

4 gels simultaneously


Electrophoresis Power Supply LA690

HiMedia Wins India's most Prestigious Rajiv Gandhi National Quality

Award Once againounded on the culture of

uncompromising quality HiMedia

have earned a global reputation

on Quality and Innovation in

BioSciences. With its gun-sight focused on quality,

this is the 2nd time HiMedia has received India’s most

prestigious Rajiv Gandhi National Quality Award. We are

happy to announce we have once again established our

Excellence in Biotechnology by receiving this award

in a glorious ceremony held in

April 2013 in New Delhi.

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