编码与大鼠ERα-甘露糖苷酶同源的人6A8cDNA在真核细胞的表达  

Expression of a human 6A8 cDNA homologous to a rat cDNA encoding an ERα mannosidase in eukaryotic cell

作　　者：张立新[1] 马凤蓉[1] 朱立平[1] 李波 石伟[1] 张淑珍[1] 李国燕[1] 王汛[1] 赵方萄[1] 

机构地区：[1]中国医学科学院基础医学研究所,中国协和医科大学基础医学院

出　　处：《中华微生物学和免疫学杂志》1998年第4期268-271,共4页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金

摘　　要：目的观察６Ａ８ｃＤＮＡ（１３５８ｂｐ）在真核细胞的表达。方法Ｎｏｒｔｈｅｒｎｂｌｏｔ，构建６Ａ８ｃＤＮＡ表达载体ｐＲｃ／ＣＭＶ－６Ａ８ｃＤＮＡ，转染ＣＯＳ－７细胞，及基因免疫小鼠。结果６Ａ８ｃＤＮＡ有４．３ｋｂ和３．５ｋｂ两种ｍＲＮＡ转录形式。４．３ｋｂｍＲＮＡ以外周血白细胞最丰富，其次为淋巴结、脾脏和骨髓，胸腺组织表达较少，胎肝则缺如。３．５ｋｂ者也以外周血白细胞最丰富，其次为肝脏、淋巴结和骨髓，胸腺和脾脏表达量最少。ｐＲｃ／ＣＭＶ－６Ａ８ｃＤＮＡ在ＣＯＳ－７细胞表达了分子量４５０００左右的蛋白质，与单抗６Ａ８有反应。ｐＲｃ／ＣＭＶ－６Ａ８ｃＤＮＡ基因免疫小鼠，６／１０只小鼠血清与人活化Ｂ细胞株３Ｄ５细胞膜有反应，５／５只对照小鼠血清呈阴性反应。结论６Ａ８ｃＤＮＡ（１３５８ｂｐ）在真核细胞获得表达，但此ｃＤＮＡ非为全长。Objective To observe the expression of 6A8 cDNA(1358bp)in eukaryotic cell. Methods Northern blot; construction of recombinant 6A8 cDNA expression vector pRc/CMV 6A8 cDNA; transfection of the expression vector into COS 7 cell; gene immunization of mice with pRc/CMV 6A8 cDNA. Results There existed two types of mRNA transcripts with molecular weight of 4.3kb and 3.5kb in various immune tissues detected with a 1358bp 6A8 cDNA as a probe. The tissues,except fetal liver,were richer in 4.3kb mRNA than in 3.5kb mRNA. The expression amount by progressive decrease for the 4.3kb mRNA was of peripheral blood leukocyte,lymph node,spleen,bone marrow and thymus. This mRNA was absent in fetal liver. For the 3.5kb mRNA peripheral blood leukocyte was also the highest followed by fetal liver,then lymph node and bone marrow,thymus and spleen.A 6A8 sense expression plasmid in an eukaryotic expression vector pRc/CMV was constructed.The 6A8 cDNA was expressed in COS 7 cell in a form of about 45kD protein which was recognized by McAb 6A8. In addition, the membrane of 3D5 cell, a human activated B cell line used for immunization in the development of the McAb,was stained by the sera from 6/10 BALB/c mice immunized with the 6A8 sense pRc/CMV expression plasmid. In contrast,the control sera of pRc/CMV injected mice were negative in reaction with 3D5 cells. Conclusion 6A8 cDNA(1358bp) can be expressed in eukaryocytes. However, this cDNA is not a full length cDNA.

关 键 词：RNA 信使 DNA 互补 甘露糖苷酶 真核细胞 

分 类 号：R392.11[医药卫生—免疫学]