PMID: 8995641Feb 1, 1997Paper

Complementation of a primer binding site-impaired murine leukemia virus-derived retroviral vector by a genetically engineered tRNA-like primer

Journal of Virology
A H LundFinn Skou Pedersen

Abstract

Reverse transcription of retroviral genomes is primed by a tRNA annealed to an 18-nucleotide primer binding site. Here, we present a primer complementation system to study molecular interaction of the replication machinery with the primer and primer binding site in vivo. Introduction of eight base substitutions into the primer binding site of a murine leukemia virus-based vector allowed efficient RNA encapsidation but resulted in severely reduced vector replication capacity. Replication was restored upon complementation with a synthetic gene designed to encode a complementary tRNA-like primer, but not with a noncomplementary tRNA-like molecule. The engineered primer was shown to be involved in both the initiation of first-strand synthesis and second-strand transfer. These results provide an in vivo demonstration that the retroviral replication machinery may recognize sequence complementarity rather than actual primer binding site and 3' primer sequences. Use of mutated primer binding site vectors replicating via engineered primers may add additional control features to retroviral gene transfer technology.

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Citations

Dec 4, 2002·Proceedings of the National Academy of Sciences of the United States of America·Mamuka KvaratskheliaStuart F J Le Grice
Jul 30, 2003·Journal of Virology·Nathan J KellyCasey D Morrow
Jan 18, 2008·Journal of Virology·Melanie GallaChristopher Baum
Nov 20, 2008·Archives of Virology·Caroline Torne-CelerCorinne Ronfort
Oct 14, 2006·The Journal of Biological Chemistry·Janine KraunusChristopher Baum
May 26, 2011·Nucleic Acids Research·Melanie GallaChristopher Baum
Oct 29, 1997·Journal of Virology·J Mak, L Kleiman

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