which sizes of dna fragments do you expect to get from the digestion?
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Journal 12.09.10:. Which sizes of DNA fragments do you expect to get from the digestion? 2. How can one use the electrophoresis procedure to estimate the size of the DNA fragments?. pARA-R construct. Recombinant plasmid of interest. pARA-R 4720 bp. pARA-R. 4720 bp. Bam H I. rfp 702bp. - PowerPoint PPT PresentationTRANSCRIPT
1. Which sizes of DNA fragments do you expect to get from the digestion?
2. How can one use the electrophoresis procedure to estimate the size of the DNA fragments?
Journal 12.09.10:
pARA-R construct
Recombinant plasmid of interest
pARA-R
BamH I
Hind III
rfp702bp
4720 bp
rfp 702bp
pARA-R4720 bp
Gel Electrophoresis - DetectionWe run the gel until the yellow dye runs out.
Wells
After running the gel (till the yellow dye runs out), we stain it in Ethidium bromide, which will fluoresce when bound to DNA:
Supercoiling
Multimers
circle “nicked-circle”
“nicked-circle”
“multimer”
Structural forms of plasmid
Linear(Digested)
Slower than nicked.
Supercoiled
10 K
b L
adder
5 Kb
MultimerNickedSuper Coiled
Linear Fragment
A- A+
Linear Fragment
10 K
b L
adder
10 K
b L
adder
M A- A+Yellow highlighter shows group number..
Expected results
Class Results 2013: Lab 4a
Groups 1,2
Groups 3,4
1000 base pairs
500 base pairs
1500 base pairs
2000base pairs4000base pairs
1) Lab 2a (restriction) and 4a (electrophoresis) almost always come together – why is that?
2) Point out at least one thing that you learned so far that is important to know when working in a biotech lab.