3GC cloning: PCR products cloning mediated by terminal deoxynucleotidyl transferase

We have developed a PCR product cloning strategy called 3GC cloning. Through this strategy, PCR products tailed with more than three homopolymeric deoxycytidines (dCs) at the 3' ends by terminal deoxynucleotidyl transferase (TdT) would anneal complementarily with three-deoxyguanosine (dG) protruding ends of 3G vector, which was generated through coupled SfiI cleavage at both recognition sites. Redundant overhangs at insert-vector junctions after ligation would be trimmed off and repaired in bacteria, and the length of such junction was only three G/C pairs. Any chain lengths of three or more nucleotides, which are able to serve as substrates for TdT, could theoretically be cloned by this strategy.