GENE REPLACEMENT OF THE P53 GENE WITH THE LACZ GENE IN MOUSE EMBRYONIC STEM-CELLS AND MICE BY USING 2 STEPS OF HOMOLOGOUS RECOMBINATION

被引:69
作者
GONDO, Y
NAKAMURA, K
NAKAO, K
SASAOKA, T
ITO, K
KIMURA, M
KATSUKI, M
机构
[1] KYUSHU UNIV 69, MED INST BIOREGULAT, FUKUOKA 812, JAPAN
[2] TOKAI UNIV, SCH MED, ISEHARA, KANAGAWA 25911, JAPAN
关键词
D O I
10.1006/bbrc.1994.2005
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Two steps of gene targeting were used to replace the p53 gene with the E.coli beta-galactosidase (lacZ) gene in mouse embryonic stem (ES) cells. The first targeting vector consisted of neo and herpes simplex virus thymidine kinase (HSV-tk) genes as a neo-tk cassette in the middle of the targeting vector. At the first targeting, the homologous recombinants became G418 resistant and ganciclovir (GANC) sensitive and were selected by G418 alone. At the second targeting, homologous recombination reciprocally exchanged the neo-tk cassette in the ES cell chromosome with the lacZ fragment in the second targeting vector and thus made the ES cells GANC resistant. We obtained two ES cell clones, in which the p53 gene for both had been replaced with a totally non-homologous sequence of the lacZ gene. The germ-line transmission of the manipulated ES cells also demonstrated that the entire procedure had no detrimental effects on ES cells at all. (C) 1994 Academic Press, Inc.
引用
收藏
页码:830 / 837
页数:8
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