dna technology chapter 13 lab biology 1a chapter 15 lab biology 2b chapter 13 honors biology 4a
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DNA
Tech
nolog
y
Chapte
r 13
Lab B
iolo
gy 1A
Chapte
r 15
Lab b
iolo
gy 2b
Chapte
r 13
Honor
s Bio
logy
4A
DNA Technology
Technology used to help with genetic engineering helps us:
1. identify genes for specific traits2. transfer genes for a specific trait from one organism to another
3. cure disease4. treat genetic disorders5. improve food crops
How can you get a desired trait without directly manipulating the organisms DNA?
1.HYBRIDIZATION; crossing organisms of different traits to produce a hardier product ex: mule
2.INBREEDING/SELECTIVE BREEDING; maintain the present genes by breeding only within the population ex: pedigree animals
3.INDUCING MUTATIONS; radiation, chemicals polyploidy (3N or 4N) plants resulted larger and hardier
Now let us manipulate the genes by altering the organisms DNA
a) DNA Technology: sci. involved in the ability to manipulate genes/DNA
a) Cure diseaseb) treat genetic disordersc) Improve crops
Tools:
1. DNA extraction
2. Restriction enzymes
3. Gel electrophoresis
4. DNA ligase
5. Polymerase chain rxn. (PCR)
Method: (5 steps)
1.Extract gene insulin2.Cut insulin producing gene out using “restriction
enzymes”1.Sticky ends create overhang2.Blunts no overhangs
3.Cutting clone vector cut plasmid with same restriction enzyme
4.Ligation: donor gene is spliced into plasmid DNA, DNA ligase glues it(this forms recombinant DNA = plasmid DNA + new piece of DNA)
5.Plasmid returned to bacterium & reproduces using donor gene in it (this is transgenic organism = organism with foreign DNA incorporated in it’s genome)
6.*reproduce*
Restriction Enzymes
BACTERIAL ENZYMES are used to cut DNA molecule into more manageable pieces
They recognize certain sequences
Creating “single-chain” tails in DNA called STICKY ENDS
Sticky Ends
Readily bind to complimentary chains of DNA therefore pieces of DNA that have been cut with the same restriction enzyme can bind together to form a new sequence of nucleotides
Recognizes CTTAAG
Cloning Vectors
Cloning vector is a carrier that is used to clone a gene and transfer it from one organism to another.
Many bacteria contain a cloning vector called a PLASMID.
PLASMID is a ring of DNA found in a bacterium in addition to its main chromosome.
PROCEDURE
To be used as a cloning vector in gene transfer experiments a plasmid is isolated from a bacterium.
Using restriction enzymes the plasmid is then cut and a DONOR GENE (specific gene isolated from another organism is spliced into it)
Then the plasmid is returned to the bacterium, where it is replicated as the bacterium divides, making copies of the donor gene.
Plasmid now contains a GENE CLONE
Transplanting genes
In some cases, plasmids are used to transfer a gene to bacteria so that the bacteria will produce a specific protein
Ex: INSULIN = protein that controls sugar metabolism
Bacteria that receives the gene for insulin will produce insulin as long as the gene is not turned off
Steps:
1. ISOLATING A GENE – isolate the DNA from human cells and plasmids from the bacteria
Use restriction enzymeSplice human DNA into plasmids to create a genomic library (set of thousands of DNA pieces from a genome that have been inserted into a cloning vector)
Steps cont…
2. PRODUCING RECOMBINANT DNA = combination of DNA from 2 or more sources
Inserting a donor gene such as human gene for insulin, into a cloning vector, such as bacterial plasmid results in a recombinant DNA molecule!
Steps cont…
3. CLONING DNA – the plasmid containing recomb. DNA is inserted into a host bacterium (called transgenic organism
The trans. Bact. Is placed in a nutrient medium where it can grow and reproduce.
Expression of Cloned Genes
Sometimes PROMOTERS must also be transferred so the genes will be turned on.
Genes are often turned off until the proteins they code for are needed.
Practical uses of DNA Technology
1. Pharmaceutical products: insulin, HBCF (human blood clotting factor)
2. Genetically engineered vaccines
3. Increased agriculteral yields
4. Improving quality of produce1.Slow down ripening2.Enhance color3.Reduce fuzz4.Increase flavor5.Frost resistance
DNA Technology Techniques
I. DNA Fingerprints pattern of bands made up of specific fragments from an individual’s DNA
USED FOR:
DETECTION OF A RELATIVESIMILARITIES BETWEEN SPECIES
How do you make DNA fingerprints?RFLP (restriction fragment length
polymorphism) analysis1. extract DNA from specimen using restriction enzymes
2. separate fragments of DNA using electrophoresis (separates DNA according to size and charge)
3. placed in wells made on gel4. electric current run through gel
Continue…5. negative fragments migrate to positive charged end of gel but not all at same rate
6. pores in gel allow smaller fragments to migrate faster separating fragments by size.
7. blotted onto filter paper.
Accuracy of DNA Fingerprints
DNA fingerprints are very accurate
However, genetic tests can only absolutely disprove, not prove, relationship!
Courts accept 99.5% accuracy as proof of alleged paternity
Polymerase Chain Reaction (PCR)
Used when you only have a TINY piece of DNA
PCR can be used to quickly make many copies of selected segments of the available DNA
Use a PRIMER to initiate replication
DNA doubles every 5 minutes
PCR is used for:1. crimes2. diagnosing genetic disorders from embryonic cells
3. studying ancient fragments of DNA (tiny amounts)
HUMAN GENOME PROJECT
2 GOALS:1. determine nucleotide
sequence of entire human genome (aprox 3 billion nucleotide pairs or about 100,000 genes
2. map the location of every gene on each chromosome
19961 % of 3 billion nucleotide
pairs of DNA human genomes were analyzed
This allows for us to identify and determine the function of 16,000 genes!
Gene TherapyTreating a genetic disorder by
introducing a gene into a cell or by correcting a gene defect in a cell’s genome.
Ex: Cyctic fibrosis cause one defective gene malfunction of one protein
Gene Therapy for Cyctic FibrosisNasal spray carrying normal
cyctic fibrosis gene to cells in nose and lungs
Must repeat treatment periodically
Ethical IssuesMany people worry about how personal genetic information will be
used:
Insurance???Employment????
Human Genome Project will undoubtedly involve ethical decisions about how society should use the information! WHAT DO YOU THINK??
Practical Uses of DNA Technology1. produce perscription drugsVaccine (harmless version of a virus or a bacterium)
Pathogen (disease causing agent) treated chemically or physically so that they can no longer cause disease.
Pathogen (Ag) Antibody (Ab)DNA tech. may produce vaccines safer than traditional ones!
Increasing Agricultural Yields
DNA Tech. used to develop new strains of plants
Ex: scientists can make tomato plants toxic to hornworms and effectively protect the plant from these pests.
Crops that do NOT need fertilizerPlants require NITROGEN to
make proteins and nucleic acidsMost plants get their N from the
soilTRANSGENIC FOOD CROPS
contain genes for nitrogen fixation so they can grow in nitrogen POOR soil.
Genetically Engineered Foods
Foods may have toxic proteins or substances causing ALLERGIES
Ex: changing the gene that codes for an enzyme to ripening in tomatoes they are able to make tomatoes ripen without becoming SOFT!!
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