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Effect of the length of 3′ overhang and the number of phosphorothioate on cloning efficiency. The mixtures of equimolar PCR fragments (10 ng pUC18 and 4 ng ape_0119) were incubated with λ exonuclease (250 ng exonuclease/pmol DNA) at 37°C for 5 min before transforming into E. coli. The primers used to generate the required overhangs are the derivatives of Ape_0119-F, Ape_0119-R, pUC18-F, and pUC18-R.
