The Sanger dideoxy method for determining DNA sequence is limited in that a stre

Question

The Sanger dideoxy method for determining DNA sequence is limited in that a stretch of only 1000 or fewer bases can be analyzed in one reaction. Suppose you wish to sequence a newly isolated double-strand DNA tumor virus that contains 5000 base pairs. You decide to use the Sanger method on restriction fragments of the DNA for sequencing. You use an enzyme that makes a significant number of cuts to give, on average, fragments of 300 nucleotides or less. Why might it be a good idea also to sequence a second set of fragments cleaved by another restriction enzyme?

Explanation / Answer

A disadvantage of Sanger method is that the sequence may contain parts of sequence of the cloning vector as well. If a second set of fragments cleaved by another restriction enzyme are taken and sequence, the possible cloning vector regions could be determined and the exact sequence could be found out.

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