Construction of pGT4ΔB
pGT4ΔB is a plasmid vector pGT4 mutant. The mutation is a disrupted BamH I restriction site. So, ΔB stands in this case for: "without BamH I site".
The construction of pGT4ΔB starts with the digestion of pGT4 with
restriction enzyme BamHI.
The experimental steps in the pGT4ΔB construction can be done simultaneously with those of the library
To create blunt ends on BamH I DNA fragments
the large fragment of E.coli DNA
polymerase is used here. This so-called Klenow enzyme is a
truncated version of polymerase I: it lacks the 5’ to 3’ exonuclease
activity. It can synthesize DNA complementary to a DNA template in a 5’
to 3’ direction, provided it is given a primer with a free 3’-OH, and