Generation of transgenic medaka using modified bacterial artificial chromosome.
The availability of bacterial artificial chromosome (BAC) offers a good genomic platform for a targeted integration of an exogenous gene by a homologous recombination system in Escherichia coli. In combination with microinjection technology, this system allows for the analysis of various aspects of biological phenomena occurring in vivo using Japanese medaka fish (Oryzias latipes). Here we describe a streamlined procedure for selecting BAC clones based on the medaka University of Tokyo genome browser (UTGB), followed by rapid modification with enhanced green fluorescent protein (EGFP) or DsRed fragments for transgenic analysis in medaka. Experimental procedures for BAC DNA preparation, microinjection of medaka embryos and screening of resulting transgenic medaka carrying EGFP/DsRed modified BAC clones are also described.
Nakamura
Shuhei
S
Laboratory of Molecular Genetics for Reproduction, National Institute for Basic Biology, Okazaki, Japan.
Saito
Daisuke
D
Tanaka
Minoru
M
D000818:Animals; D030801:Animals, Genetically Modified; D022202:Chromosomes, Artificial, Bacterial; D015509:Developmental Biology / Q000379:methods; D004926:Escherichia coli; D005821:Genetic Techniques; D016678:Genome; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008957:Models, Genetic; D009990:Oryzias / Q000235:genetics*; D011995:Recombination, Genetic; D019076:Transgenes
D049452:Green Fluorescent Proteins