血液系统条件性DNMT 3B 基因敲除小鼠的构建及鉴定

Generation of hematopoietic system-specific DNMT3B gene conditional knockout in mouse model

摘要:

目的 构建可在血液系统中条件性敲除DNMT 3B 基因小鼠并进行鉴定，为研究DNMT 3B 基因的生物学功能及其在血液肿瘤疾病的作用机制提供动物模型。方法 将引进的DNMT3Bflox/flox 小鼠与Mx1-Cre+ 小鼠进行杂交繁殖，得到基因型DNMT3Bflox/+Mx1-Cre+ 及DNMT3Bflox/+Mx1-Cre- 两种子代，再用这两种子代杂交产生子代小鼠，通过PCR 法鉴定子代小鼠的基因型；获得基因型为DNMT3Bflox/flox Mx1-Cre+ 的小鼠，通过腹腔注射pI-pC 共5 次诱导DNMT3B 敲除，然后用半定量PCR 方法鉴定DNMT 3B 基因敲除情况。结果 基因型鉴定确认在13 只子鼠中有2 只为DNMT3Bflox/flox Mx1-Cre+，经pI-pC 腹腔注射后，DNMT 3B 基因通过Cre/loxP 系统被成功敲除。结论 该方法成功构建可以在血液系统条件性敲除DNMT 3B 基因的小鼠，并进行敲除后鉴定。

Abstract:

Objective To generate hematopoietic system-specific DNMT3B conditional knockout mice,providing mouse model for further functional study of DNMT3B in hematological malignancies. Methods DNMT3Bflox/flox mice were cross bred with Mx1-Cre+ mice to generate DNMT3Bflox/+ Mx1-Cre+ and DNMT3Bflox/+ Mx1- Cre- mice. DNMT3Bflox/+ Mx1-Cre+ mice were further cross bred with DNMT3Bflox/+ Mx1-Cre- mice for DNMT3Bflox/flox Mx1-Cre+ mice. DNMT3Bflox/flox Mx1-Cre+ mice were injected intraperitoneally with poly (I:C) twice a day for a total of five times for DNMT3B deletion. PCR was performed for genotyping of off-springs. Deletion of DNMT3B was confirmed by semi-quantitative PCR. Results Genotyping results identified 2 out of 13 off-springs which were DNMT3Bflox/flox Mx1-Cre+ homozygous. DNMT3B gene was successfully knocked out with injection of pI-pC.Conclusions Hematopoietic system-specific DNMT3B gene conditional knockout homozygous mice are successfully generated, which serves as an ideal model for functional study of DNMT3B.