Question

Question 1. Consider the segment of DNA below, taken from several CCNY rattus. From each rat, the fragment was amplified using the primers shown. Some of the amplification product was immediately run on a DNA agarose gel: the remainder was treated with EcoRI, then run on a separate gel. On the gel representation below, show where each DNA fragment would migrate for the individuals described: A. homozygous wildtype, before EcoRI treatment B. homozygous mutant, before EcoRI treatment C. heterozygous, before EcoRI treatment D. homozygous wildtype, after EcoRI treatment E. homozygous mutant, after EcoRI treatment F. heterozygous, after EcoRI treatment before EcoRI after EcoRI treatment treatment

Can someone explain this, I do not understand, I have the answer but I just don't understand why?

Answer 1

before EcoRI after EcoRI treatment treatment

Bands are colour-coded for easy analysis.

In lanes

A: Full-length fragment will be formed

B: Shorter than A, because of the deletion of the region

C: Will have both bands of A and B, as it is heterozygous for wt and mutant

D: Has three EcoRI sites, if the sites are included in the primers, otherwise only one EcoRI site in the center. In any case, the EcoRI site in the center one will give one cut in the PCR product and will have two bands. The sites on the end will have no effect on product size.

E: Has the EcoRI site region deleted in the product. So only full length will appear. No effect of EcoRI treatment.

F: Will be a mix of two products, will give three bands. One corresponding to the full length of mutant DNA. Two from the wt product with EcoRI site in the middle.