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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.

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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

BamH I

rfp

702bp

rfp 702bp

Hind III


Gel Electrophoresis - Detection

We 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 stain it in Ethidium bromide, which will fluoresce when bound to DNA:

Multimers

“nicked-circle”

circle


Structural forms of plasmid stain it in Ethidium bromide, which will fluoresce when bound to DNA:

Slower than nicked.

“multimer”

“nicked-circle”

Linear

(Digested)

Supercoiled


A- stain it in Ethidium bromide, which will fluoresce when bound to DNA:

A+

10 Kb Ladder

10 Kb Ladder

10 Kb Ladder

Multimer

Nicked

Super Coiled

5 Kb

Linear Fragment

Linear Fragment


M stain it in Ethidium bromide, which will fluoresce when bound to DNA:

A-

A+

Yellow highlighter shows group number..

Expected results


Class Results 2013: Lab 4a stain it in Ethidium bromide, which will fluoresce when bound to DNA:


Groups 1,2 stain it in Ethidium bromide, which will fluoresce when bound to DNA:


Groups 3,4 stain it in Ethidium bromide, which will fluoresce when bound to DNA:


4000base pairs stain it in Ethidium bromide, which will fluoresce when bound to DNA:

2000base pairs

1500 base pairs

1000 base pairs

500 base pairs



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