人杀伤抑制性受体CD158b cDNA的克隆及其在大肠杆菌中的表达  

Cloning of human natural killer cell inhibitory receptor CD158b cDNA and its expression in E.coli

作　　者：曹东林[1] 周思朗[2] 白英明[1] 何肖娟[1] 

机构地区：[1]第一军医大学分校生化教研室,广东省广州市510315 [2]海军421医院老干病房,广东省广州市510318

出　　处：《第一军医大学分校学报》2004年第1期10-12,共3页Journal of Branch Campus of the First Military Medical University

基　　金：国家自然科学基金 (39870 330 ) ;广东省自然科学基金 (0 0 1 0 4 3)

摘　　要：目的 :CD1 5 8b基因克隆与表达。方法 :采用RT -PCR技术 ,从正常人外周血单个核细胞中扩增编码CD1 5 8b的cDNA ,经酶切后将其克隆于pMBP -c表达质粒上 ,酶切和测序鉴定。构建的高效表达克隆子经IPTG诱导后 ,表达的重组蛋白经SDS -PAGE电泳分析。结果 :RT -PCR获得了预期大小的cDNA ,DNA测序结果与GenBank登记的人CD1 5 8b编码序列一致。pMBP -c -CD1 5 8b表达质粒酶切后结果与预期相符。表达的重组蛋白经SDS -PAGE电泳分析 ,得到分子量为 5 8kD的蛋白质 ,与理论值一致 ,所获重组蛋白占菌体总蛋白 2 5 %。结论 :获得了CD1 5 8b基因 ,并成功在大肠杆菌中表达 ,所获重组蛋白为进一步研究CD1 5Objective To clone and express the CD158b gene.Methods The gene which encodes matural killer cell inhibitory receptor was amplified by RT-PCR from peripheral blood mononuclear cells(PBMCs). Recombinant expression vector was constructed and sequenced after enzyme digestion. Results (1)Prospective amplified cDNA was obtained and the base sequence was identical with that recorded in GenBank after DNA sequencing. (2)Identification of pMBP-c-CD158b expression vector showed prospective result after the enzyme digestion analysis. (3)The size of recombinant protein was also coincident with the anticipotive aim and its proportion was 25% of the total protein. Conclusion CD158b cDNA was obtained by RT-PCR and successfully expressed in E.coli . The recombinant protein could provide an essential preparation for studying its functions.

关 键 词：CDl58b基因 逆转录聚合酶链式反应 表达质粒 造血干细胞移植 

分 类 号：R457.7[医药卫生—治疗学]