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Knockout mouse production assisted by Blm knockdown
Authors:Mikiko FUKUDA  Mayuko INOUE  Daisuke MURAMATSU  Hitoshi MIYACHI  Yoichi SHINKAI
Institution:1)Cellular Memory Laboratory, RIKEN, Saitama 351-0198, Japan;2)CREST (Core Research for Evolutionary Science and Technology), Japan Agency for Medical Research and Development (AMED), Japan;3)Institute for Virus Research, Kyoto University, Kyoto 606-8507, Japan;4)Graduate School of Biostudies, Kyoto University, Kyoto 606-8501, Japan
Abstract:Production of knockout mice using targeted embryonic stem cells (ESCs) is a powerful approach for investigating the function of specific genes in vivo. Although the protocol for gene targeting via homologous recombination (HR) in ESCs is already well established, the targeting efficiency varies at different target loci and is sometimes too low. It is known that knockdown of the Bloom syndrome gene, BLM, enhances HR-mediated gene targeting efficiencies in various cell lines. However, it has not yet been investigated whether this approach in ESCs is applicable for successful knockout mouse production. Therefore, we attempted to answer this question. Consistent with previous reports, Blm knockdown enhanced gene targeting efficiencies for three gene loci that we examined by 2.3–4.1-fold. Furthermore, the targeted ESC clones generated good chimeras and were successful in germline transmission. These data suggest that Blm knockdown provides a general benefit for efficient ESC-based and HR-mediated knockout mouse production.
Keywords:Blm  Embryonic stem cells (ESCs)  Gene targeting
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