Template-directed chemical ligation to obtain 3'-3' and 5'-5' phosphodiester DNA linkages.

Abstract

Up to now, the direct ligation of two DNA fragments with opposite directions to obtain 3′-3′ or 5′-5′ phosphate ester bonds is still challenging. The only way to obtain DNA oligonucleotides containing a 3′-3′ or 5′-5′ inversion of polarity sites is based on professional DNA chemical synthesis. Herein, we demonstrate a convenient template-directed chemical ligation that enables 3′-3′ and 5′-5′ linkages of two DNA oligonucleotides. This method is based on the assembly of two oligonucleotides on a template in opposite directions through forming antiparallel and parallel duplexes simultaneously, followed by coupling with N-Cyanoimidazole under mild condition. Moreover, on the basis of DNA oligonucleotides with 5′-5′ linkage obtained through our template-directed chemical ligation, we developed a new cDNA display technique for in vitro selection of functional polypeptides.