Chapter 9, Problem 16P

Suppose you are using a plasmid cloning vector that has no EcoRI sites (5′ G^AATTC 3′) in its polylinker because the particular drug resistance gene your vector contains has an EcoRI site within it.

a.	How could you use the following two oligonucleotides (and ligase enzyme) to ligate an insert that is an EcoRI fragment into the BamHI site (5′ G^GATCC 3′) in the polylinker of your vector?
b.	How many EcoRI sites will the recombinant DNA contain? How many BamHI sites?
c.	In part (a), you used the two oligonucleotides to make a so-called adapter. Adapters can also be used to ligate blunt-ended inserts into vectors cut with sticky-ended enzymes. Design an adapter that would allow you to ligate blunt-ended inserts into the BamHI site of your vector’s polylinker. (Note: Two blunt-ended DNA fragments can be ligated together, although the reaction is much less efficient than sticky-end ligation.)