Experimental Animals
Online ISSN : 1881-7122
Print ISSN : 1341-1357
ISSN-L : 0007-5124
Original
Establishment of an Efficient BAC Transgenesis Protocol and its Application to Functional Characterization of the Mouse Brachyury Locus
Kuniya ABEMasaaki HAZAMAHideki KATOHKen-ichi YAMAMURAMisao SUZUKI
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JOURNAL FREE ACCESS

2004 Volume 53 Issue 4 Pages 311-320

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Abstract

Transgenesis using large DNA such as YAC or BAC has extended the range of applications in functional genomics. Here we describe an efficient BAC transgenesis protocol using a simple BAC DNA preparation method adopted from YAC DNA purification methods. This method allowed us to isolate BAC DNA from small scale culture of BAC-containing cells in sufficient quantity and purity for microinjection. More than 40 founders have been produced with linearized BAC DNA prepared by this method, and 85% of them contained intact BAC transgenes. In contrast, when circular BAC DNA was injected, an approximately three-fold reduction of transgene integration rate was observed and fewer intact transgene integrations were obtained. A line of transgenic mice carrying a 170-kb BAC clone generated in this way successfully rescued tail and embryonic lethality phenotypes of the mouse Brachyury (T) mutants, further demonstrating the utility of this method in functional analysis of the mouse genome.

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© 2004 Japanese Association for Laboratory Animal Science
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