title: In vivo site-specific integration of transgene in silkworm via PhiC31 integrase-mediated cassette exchange.
abstract: Current techniques for genetic engineering of the silkworm Bombyx mori genome utilize transposable elements, which result in positional effects and insertional mutagenesis through random insertion of exogenous DNA. New methods for introducing transgenes at specific positions are therefore needed to overcome the limitations of transposon-based strategies. Although site-specific recombination systems have proven powerful tools for genome manipulation in many organisms, their use has not yet been well established for the integration of transgenes in the silkworm. We describe a method for integrating target genes at pre-defined chromosomal sites in the silkworm via phiC31/att site-specific recombination system-mediated cassette exchange. Successful recombinase-mediated cassette exchange (RMCE) was observed in the two transgenic target strains with an estimated transformation efficiency of 3.84-7.01%. Our results suggest that RMCE events between chromosomal attP/attP target sites and incoming attB/attB sites were more frequent than those in the reciprocal direction. This is the first report of in vivo RMCE via phiC31 integrase in the silkworm, and thus represents a key step toward establishing genome manipulation technologies in silkworms and other lepidopteran species.
authors: State Key Laboratory of Silkworm Genome Biology, Key Laboratory for Sericulture Functional Genomics and Biotechnology of Agricultural Ministry, Southwest University, BeiBei, Chongqing 400716, China.
authors: Dingpei
authors: D
authors: Long
authors: Aichun
authors: A
authors: Zhao
authors: Longxia
authors: L
authors: Xu
authors: Weijian
authors: W
authors: Lu
authors: Qing
authors: Q
authors: Guo
authors: Yang
authors: Y
authors: Zhang
authors: Zhonghuai
authors: Z
authors: Xiang
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics*; D001435:Bacteriophages / Q000201:enzymology*; D012831:Bombyx / Q000235:genetics*; D019426:Integrases / Q000235:genetics / Q000378:metabolism*; D016254:Mutagenesis, Insertional / Q000379:methods*; D011995:Recombination, Genetic; D019076:Transgenes; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D014764:Viral Proteins; D019426:Integrases
keywords: Bombyx mori; Site-specific integration; Transgenic silkworm; phiC31/att system; piggyBac