The restriction endonuclease cleavage sites for SphI and KpnI have been added to the lac cloning region of the phage vectors M 13mp10 and M 13mp11, using oligodeoxynucleotide-directed in vitro mutagenesis. Complementary deoxy 16-, 21- or 18-mers with the desired base changes were annealed to the M13mp DNA strand and extended with the Klenow fragment of DNA polymerase I. In adding these sites we have shown that this technique can be used as a general method for inserting sequences of DNA as well as introducing deletions and base pair changes.
- DNA polymerase I Klenow fragment
- M13 cloning
- gene synthesis
- synthetic DNA primers