Modified bacterial artificial chromosomes for zebrafish transgenesis.
Transgenesis using bacterial artificial chromosomes (BAC) offers greater fidelity in directing desirable expression of foreign genes. Application of this technology in the optically transparent zebrafish with fluorescent protein reporters enables unparalleled visual analysis of regulation of gene expression in a living organism. Here we describe a streamlined procedure of direct selecting multiple BAC clones based on public sequence databases followed by rapid modification with GFP or RFP for transgenic analysis in zebrafish. Experimental procedures for BAC DNA preparation, microinjection of zebrafish embryos and screening of transgenic zebrafish carrying GFP/RFP modified BAC clones are detailed.
Yang
Zhongan
Z
Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, CA 90095-1606, USA.
Jiang
Hong
H
Chaichanasakul
Thawinee
T
Chachainasakul
Thawinee
T
Gong
Shiaoching
S
Yang
Xiangdong William
XW
Heintz
Nathaniel
N
Lin
Shuo
S
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics*; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D004625:Embryo, Nonmammalian; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D017930:Genes, Reporter; D005818:Genetic Engineering / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000032:analysis; D008164:Luminescent Proteins / Q000032:analysis; D010957:Plasmids / Q000235:genetics; D011938:Rec A Recombinases / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000032:analysis; D011995:Recombination, Genetic; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*; D000097573:Red Fluorescent Protein
D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D011938:Rec A Recombinases; D011993:Recombinant Fusion Proteins; D000097573:Red Fluorescent Protein