In vivo site-specific integration of transgene in silkworm via PhiC31 integrase-mediated cassette exchange. 2013

Dingpei Long, and Aichun Zhao, and Longxia Xu, and Weijian Lu, and Qing Guo, and Yang Zhang, and Zhonghuai Xiang
State Key Laboratory of Silkworm Genome Biology, Key Laboratory for Sericulture Functional Genomics and Biotechnology of Agricultural Ministry, Southwest University, BeiBei, Chongqing 400716, China.

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.

UI MeSH Term Description Entries
D011995 Recombination, Genetic Production of new arrangements of DNA by various mechanisms such as assortment and segregation, CROSSING OVER; GENE CONVERSION; GENETIC TRANSFORMATION; GENETIC CONJUGATION; GENETIC TRANSDUCTION; or mixed infection of viruses. Genetic Recombination,Recombination,Genetic Recombinations,Recombinations,Recombinations, Genetic
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D001435 Bacteriophages Viruses whose hosts are bacterial cells. Phages,Bacteriophage,Phage
D012831 Bombyx A genus of silkworm MOTHS in the family Bombycidae of the order LEPIDOPTERA. The family contains a single species, Bombyx mori from the Greek for silkworm + mulberry tree (on which it feeds). A native of Asia, it is sometimes reared in this country. It has long been raised for its SILK and after centuries of domestication it probably does not exist in nature. It is used extensively in experimental GENETICS. (From Borror et al., An Introduction to the Study of Insects, 4th ed, p519) Bombyx mori,Silkmoths,Silkworms,Silkmoth,Silkworm,Bombyx morus,Bombyxs,mori, Bombyx
D014764 Viral Proteins Proteins found in any species of virus. Gene Products, Viral,Viral Gene Products,Viral Gene Proteins,Viral Protein,Protein, Viral,Proteins, Viral
D016254 Mutagenesis, Insertional Mutagenesis where the mutation is caused by the introduction of foreign DNA sequences into a gene or extragenic sequence. This may occur spontaneously in vivo or be experimentally induced in vivo or in vitro. Proviral DNA insertions into or adjacent to a cellular proto-oncogene can interrupt GENETIC TRANSLATION of the coding sequences or interfere with recognition of regulatory elements and cause unregulated expression of the proto-oncogene resulting in tumor formation. Gene Insertion,Insertion Mutation,Insertional Activation,Insertional Mutagenesis,Linker-Insertion Mutagenesis,Mutagenesis, Cassette,Sequence Insertion,Viral Insertional Mutagenesis,Activation, Insertional,Activations, Insertional,Cassette Mutagenesis,Gene Insertions,Insertion Mutations,Insertion, Gene,Insertion, Sequence,Insertional Activations,Insertional Mutagenesis, Viral,Insertions, Gene,Insertions, Sequence,Linker Insertion Mutagenesis,Mutagenesis, Linker-Insertion,Mutagenesis, Viral Insertional,Mutation, Insertion,Mutations, Insertion,Sequence Insertions
D019076 Transgenes Genes that are introduced into an organism using GENE TRANSFER TECHNIQUES. Recombinant Transgenes,Recombinant Transgene,Transgene,Transgene, Recombinant,Transgenes, Recombinant
D019426 Integrases Recombinases that insert exogenous DNA into the host genome. Examples include proteins encoded by the POL GENE of RETROVIRIDAE and also by temperate BACTERIOPHAGES, the best known being BACTERIOPHAGE LAMBDA. Integrase
D030801 Animals, Genetically Modified ANIMALS whose GENOME has been altered by GENETIC ENGINEERING, or their offspring. Animals, Transgenic,Genetically Modified Animals,Transgenic Animals,Founder Animals, Transgenic,GMO Animals,Genetically Engineered Animals,Animal, GMO,Animal, Genetically Engineered,Animal, Genetically Modified,Animal, Transgenic,Animal, Transgenic Founder,Animals, GMO,Animals, Genetically Engineered,Animals, Transgenic Founder,Engineered Animal, Genetically,Engineered Animals, Genetically,Founder Animal, Transgenic,GMO Animal,Genetically Engineered Animal,Genetically Modified Animal,Modified Animal, Genetically,Modified Animals, Genetically,Transgenic Animal,Transgenic Founder Animal,Transgenic Founder Animals

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