基于piggyBac转座子的粟酒裂殖酵母插入诱变技术

piggyBac transposon-based insertional mutagenesis for the fission yeast Schizosaccharomyces pombe.

作者信息

Li Jun, Du Li-Lin

机构信息

National Institute of Biological Sciences, 7 Science Park Road, Zhongguancun Life Science Park, Beijing, 102206, People's Republic of China.

出版信息

Methods Mol Biol. 2014;1163:213-22. doi: 10.1007/978-1-4939-0799-1_16.

DOI:10.1007/978-1-4939-0799-1_16

PMID:24841310

Abstract

Transposon-mediated insertional mutagenesis is a powerful tool for genetic screens. We have developed a piggyBac transposon-based mutagenesis system for the fission yeast Schizosaccharomyces pombe. Here, we describe in detail the procedure for inducing and selecting transpositions, and two protocols for identifying the transposon insertion sites, one using inverse PCR to identify the insertion sites in individual mutants, and the other using high-throughput sequencing to reveal the insertion sites in a mutant pool containing hundreds to thousands of mutants.

摘要

转座子介导的插入诱变是一种用于基因筛选的强大工具。我们已经为裂殖酵母粟酒裂殖酵母开发了一种基于piggyBac转座子的诱变系统。在这里，我们详细描述了诱导和选择转座的程序，以及两种鉴定转座子插入位点的方案，一种使用反向PCR来鉴定单个突变体中的插入位点，另一种使用高通量测序来揭示包含数百至数千个突变体的突变体库中的插入位点。