Question

14. Like a ladder. Circular DNA from the SV40 virus was isolated and subjected to gel electrophoresis. The results are shown in lane A (the control) of the adjoining gel patterns.

(a) Why does the DNA separate in agarose gel electrophoresis? How does the DNA in each band differ?

(b) What types of DNA do the various bands represent?

(c) What is the significance of the fact that more of the DNA is in slower-moving forms?

Answer 1

1. Agarose gel has some pore size which is suitable for DNA size range. DNA is negatively charged and moves towards positive side of electrode when gel is place between positive and negative side of electrode. During this movement DNA of bigger size moves slowly due of restriction of pore size while small size moves faster. DNA in each band differ by size

2. Different size DNA in circular DNA sample can be attributed to supercoiling. Double stranded circular DNA can be supercoiled by two ways: writhe and twist the overall sum of both is called linking number. High linking number represents the lower size of DNA just like between two similar umbrella open umbrella shows big 3D size while closed one having small 3D size.

3. Size and distance travelled in gel has negative logarithmic relation means size reduces 10 time but distance travelled increases by 2 times. Here virus DNA isolated was run in the gel and its different size in agarose gel can be attributed to super coiling and broken DNA due to some stress while isolating the DNA. Supercoiling changes the 3D size of circular DNA but not drastic so most of DNA is in slow moving forms in gel