You use the primer 5\' GCCTCGAATCGGGTACC 3\' to sequence part of a human DNA ins

Question

You use the primer 5' GCCTCGAATCGGGTACC 3' to sequence part of a human DNA insert of a recombinant DNA molecule made with a plasmid vector. The result of the automated Sanger sequencing analysis is shown below. Write the sequence of all the nucleotides of human DNA that you can determine. Indicate the 5' to 3' orientation of this sequence. Is the sequence from (a) part of the new DNA strand that was synthesized in the sequencing reaction or part of the template strand used in the sequencing reaction? Why? If you were performing this experiment, how would you know how to design the appropriate primer for the sequencing reaction? Draw the recombinant DNA molecule that was sequenced. Indicate the human and vector sequences, the position and orientation of the primer, and the position and orientation of new DNA synthesized during the sequencing reaction. (See Fig. 9.7.) Give the full sequence of the smallest DNA molecule synthesized in the sequencing reaction that contains a dideoxyG (ddG). Indicate the 5'-to-3' orientation of the molecule and the location of the ddG.

Explanation / Answer

A) 5- TTTGCTTTGTGAGCGGATAACAA.-3.. This is the sequence of the new strand which is developed.!! It's complementary sequence will be the sequence of human DNA 3'- AAACAAACACTCGCCTATTGTT- 5'..

B) the above sequence is the sequence of the new starnd which is being formed because the nucleotide are given colour when the new nucleotide are incorporated I gives the color band in the computer..!!

C) first I will sequence the the new strand sequence.. Then I can determine th sequence of the old strand then through this we can design a primer..!!

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