Directional cloning of native PCR products with preformed sticky ends (Autosticky PCR) |
| |
Authors: | J Gál R Schnell S Szekeres M Kálmán |
| |
Institution: | Institute for Biotechnology Bay Zoltán Foundation for Applied Research H-6701 Szeged, P.O. Box 2337, Hungary e-mail: gal@bay.u-szeged.hu Tel.: +36-62-432251, Fax: +36-62-432250, HU
|
| |
Abstract: | A novel method for the directional cloning of native PCR products was developed. Abasic sites in DNA templates make DNA polymerases stall at the site during synthesis of the complementary strand. Since the 5′ ends of PCR product strands contain built-in amplification primers, abasic sites within the primers result in the formation of 5′ single-stranded overhangs at the ends of the PCR product, enabling its direct ligation to a suitably cleaved cloning vector without any further modification. This “autosticky PCR” (AS-PCR) overcomes the problems caused by end sensitivity of restriction enzymes, or internal restriction sites within the amplified sequences, and enables the generation of essentially any desired 5′ overhang. |
| |
Keywords: | |
本文献已被 SpringerLink 等数据库收录! |
|