BALB/c小鼠胚胎干细胞系建立的方法学探讨

CORC > 北京大学 > 生命科学学院

	BALB/c小鼠胚胎干细胞系建立的方法学探讨; Researches on the Methodology of Establishment of BALB/c Murine Embryonic Stem Cell Lines
	孟国良 ; 滕路 ; 薛友纺 ; 尚克刚
刊名	遗传学报
	2002
关键词	BALB/c小鼠胚胎干细胞大鼠心脏细胞条件培养基(RH-CM) 内细胞团(ICM) 二倍体核型
英文摘要	以小鼠胚胎成纤维细胞(ME)为饲养层,以大鼠心脏细胞条件培养基(RH-CM)为ES细胞培养基,全面、详尽地对BALB/c小鼠ES细胞的建系和培养方法进行了探讨,成功地建立了一套建立和培养BALB/c小鼠ES细胞系的新方法.这一培养条件不但有效地维持了ES细胞的未分化状态和正常二倍体核型,而且维持了其作为多能性胚胎干细胞的一系列特征;实验设计了两种离散方法和两种浓度的消化液,用来离散增殖的ICM和ICM离散后出现的ES集落.两种离散方法即"一次离散法",和"多次离散法",两种浓度的消化液即0.25髎 Trypsin-0.04髎 EDTA和0.05髎 Trypsin-0.008髎 EDTA;同时对ICM离散时机、RH-CM在BALB/c小鼠ES细胞建系和培养中作用进行了探讨.结果表明:在低浓度消化液作用下,采用"多次离散法"离散增殖4天的ICM和ES集落的方法建立BALB/c小鼠的ES细胞系是理想的;从细胞形态、集落形态、增殖生长能力、核型检测、碱性磷酸酶测定以及体内外分化能力表明,所建立的9个BALB/c小鼠ES细胞系符合小鼠胚胎干细胞的一系列特征.; Several methods and processes of establishment and culture of BALB/c mouse ES cell lines were discussed detailedly. A new method to establish and culture ES cell lines derived from BALB/c mouse was set up successfully using mouse embryonic fibroblast feeder layer and rat-heart-cell-conditioned medium (RH-CM). These culture conditions not only maintain the undifferentiated state and normal diploid karyotypes of BALB/c mouse ES cells effectively, but also maintain a series of their characteristics of murine stem cells. two different kinds of digestive methods and Two kinds of digestive juice with different concentrations were designed to dissociate proliferous inner cell mass (ICM) and ES cell colonies derived from dissociative ICM. two different kinds of digestive methods are "single time dissociation method" and "several times dissociation method", two kinds of digestive juice are 0.25% Trypsin-0.04% EDTA and 0.05% Trypsin-0.008% EDTA. At the same time, appropriate dissociated occasion of ICM and the effect of RH-CM on establishment and culture of BALB/c mouse ES cell lines were discussed. The results suggested that it is a reasonable method to establish BALB/c mouse ES cell lines using low concentration digestive juice and "several times dissociation method" to dissociate ICM after 4 days' proliferation. Judged by the form of ES cells and its colonies, proliferous capability, karyotypes examine, alkaline phosphatase activity assay and differentiation capability in vitro and in vivo, the 9 ES cell lines that we established satisfied the all traits of murine ES cell line.; 国家自然科学基金; PubMed; 中文核心期刊要目总览(PKU); 中国科学引文数据库(CSCD); 0; 7; 581-588; 29
语种	中文
内容类型	期刊论文
源URL	[http://ir.pku.edu.cn/handle/20.500.11897/45614]
专题	生命科学学院
推荐引用方式 GB/T 7714	孟国良,滕路,薛友纺,等. BALB/c小鼠胚胎干细胞系建立的方法学探讨, Researches on the Methodology of Establishment of BALB/c Murine Embryonic Stem Cell Lines[J]. 遗传学报,2002.
APA	孟国良,滕路,薛友纺,&尚克刚.(2002).BALB/c小鼠胚胎干细胞系建立的方法学探讨.遗传学报.
MLA	孟国良,et al."BALB/c小鼠胚胎干细胞系建立的方法学探讨".遗传学报 (2002).