血红素加氧酶-1显性负性突变体转基因小鼠模型的建立与鉴定  

Establishment of a Transgenic Mouse Model of Heme Oxygenase-1 Dominant Negative Mutation

作　　者：王波[1] 程萱[2] 刘庆军[1] 魏旭冉[1] 尹玉静[1] 高旭[3] 杨晓[2] 周虹[1] 

机构地区：[1]军事医学科学院野战输血研究所,北京100850 [2]军事医学科学院生物工程研究所,北京100071 [3]哈尔滨医科大学生化教研室,哈尔滨150081

出　　处：《中国实验动物学报》2010年第3期181-184,I0003,共5页Acta Laboratorium Animalis Scientia Sinica

基　　金：国家973项目(2009CB521702)

摘　　要：目的建立血红素加氧酶-1(heme oxygenase-1,HO-1)显性负性突变体G143H转基因小鼠模型。方法通过SalI/DraI酶切pCAGGHO-1G143H转基因表达载体,纯化回收HO-1G143H表达盒片段;通过显微注射把表达目的基因的DNA片段导入FVB小鼠受精卵原核,并移植给同期发情的假孕受体母鼠,获得子代小鼠;用PCR对子代鼠尾DNA进行鉴定,并用Southern blot对结果做进一步验证;通过RT-PCR、免疫组化和Western blot方法检测HO-1基因的表达。结果表达盒回收片段正确;假孕鼠出生的17只子代小鼠共有3只阳性,均为雄性;RT-PCR、免疫组化和Western blot结果表明,阳性小鼠体内的HO-1 mRNA与蛋白表达水平增高。结论成功建立HO-1显性负性突变体G143H表达的转基因小鼠,该模型为研究HO-1在体内的作用机制奠定了基础。Objective To establish a HO-1 dominant negative mutant G143H transgenic mouse model.Methods The purified HO-1G143H fragment,digested from pCAGGHO-1G143H plasmid with SalI and DraI,was microinjected into FVB superovulated pronuclear zygotes.The injected zygotes were transplanted into the oviducts of pseudopregnant mice.The genotype of transgenic mice was identified by PCR and Southern blot.The HO-1 expression in the tissues of the transgenic mice was detected by RT-PCR,immunohistochemistry and Western blot.Results Three positive transgenic male mice were detected from 17 viable offsprings.RT-PCR,immunohistochemistry and Western blot results demonstrated that the levels of HO-1 mRNA and protein were increased in the positive mice.Conclusion A HO-1 dominant negative mutant HO-1G143H transgenic mouse model has been established successfully.The transgenic mice may serve as a suitable model for the research of HO-1action mechanisms in vivo.

关 键 词：HO-1 显微注射法 转基因小鼠 

分 类 号：Q3[生物学—遗传学]