Evaluating the Role of G,C-nucleotides and Length of Overhangs in T4 DNA Ligase Efficiency

Abstract

Ligation of fragments generated by different restriction enzymes has been observed to show variation in efficiency. In this study various restriction enzymes have been investigated to determine whether G,C-nucleotide content and/or overhang length may be contributing factors in ligation of pUC19 by T4 DNA ligase. Results suggest that G,C-rich cohesive ends may help to improve ligation efficiency. In the case of overhangs containing A,G,T,C nucleotides, it may be beneficial to use longer overhangs than shorter ones. A,Trich overhangs appear to correspond to poor ligation. Choosing appropriate termini may help to improve ligation efficiency.