To perform a DNA sequencing reaction, in each of the four reaction tubes you add

Question

To perform a DNA sequencing reaction, in each of the four reaction tubes you added "P-labeled primer, Taq DNA polymerase, DNA template, buffer, dNTP mix, and one of the four ddNTPs. After the reaction four reaction products were run in four separate lanes of a polyacrylamide gel as shown below. The ddNTP used in a reaction is indicated on top of the gel. Based on the banding pattern answer the questions listed below. For lanes B refer to the questions B. Q1 ddATP ddTTP ddCTP ddGTP B Lane 1 2 Write the sequence of the first four bases (reading from the 3'-end) of the template DNA strand. Mark the 3'- end of the sequence. 2 pts A) Suppose a sequencing reaction was prepared which contained all four dNTPs but no ddNTP. If the products of this reaction were run in lane 5 of the gel shown above, what banding pattern would be expected? Draw the banding pattern on the gel above and give a reason in one or two sentences in support of your answer. 2 pt B)

Explanation / Answer

the sequence according to me would be complementary to the shown sequence HENCE, 3'-CATT (first 4 nucleotide)

if no ddNTP is added, a single band would be observed at the end of the lane like in case of last band of ddATP as the replication of DNA would not stop and polymerase would keep on adding all nucleotide till the last template nucleotide.

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