小鼠Cdc25B融合蛋白构建和表达  

Consturction and expression of mouse Cdc25B protein

作　　者：赵鸿梅[1] 滕秋艳[2] 张哲[1] 于秉治[1] 

机构地区：[1]辽宁省人民医院检验科,沈阳110016 [2]中国医科大学生化与分子生物学教研室

出　　处：《中国公共卫生》2007年第8期976-977,共2页Chinese Journal of Public Health

基　　金：教育部高等学校博士学科点专项科研基金资助(20050159019)

摘　　要：目的研究小鼠N末端缺失286氨基酸的野生型Cdc25B原核表达载体的构建和表达。方法应用Primer5.0软件分析并设计小鼠Cdc25B5′及3′端引物(N末端缺失286氨基酶),化学合成并用聚合酶链反就(PCR)方法扩增,经限制性内切酶酶切后连接原核表达载体PGEX4T-2,构建成PGEX4T-2-Cdc25B融合表达载体,转化大肠埃希菌BL21后,通过异丙基硫代-β-D半乳糖苷(IPTG)诱导进行目的蛋白表达。结果通过双酶切、电泳分析及测序证明成功构建了Cdc25B融合表达载体,融合蛋白产物经蛋白印迹(Westernblot)显示,相对分子量为53KD的蛋白谱带,与预期一致。结论成功构建并表达了Cdc25B-N末端缺失的基因,并在原核细胞中获得较高水平的表达,为近一步研究Cdc25B的功能提供基础依据。Objective To study construction and expression of mouse cde25B protein. Methods The Cdc2.5B gene was amplified by PCR, enzyme digest and linked behind PGEX4T-2. After inducing with IPTG, the target gene was primarily ex- pressed. Results The CAc2.5B gene consisted of about 900bp by DNA eleetrophoresis. The prokaryotic expression vector of PGEX4T-2/CAc25B was constructed. After induction, a new anticipated Mr 53KD band appeared in Westen-Blot. Conclusion Fusion proteins of GST/Cdc25B are successfully expressed in BL21. This study is a basis for the role of Cdc25B during mouse oocyte maturatin.

关 键 词：CDC25B 基因表达 谷胱甘肽转移酶(GST) 聚合酶链式反应 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]