人结肠癌组织环加氧酶-2 mRNA剪接异构体片段分离与克隆  

Isolation and Cloning of Cyclooxygenase-2 Alternative Splice Variant in Human Colon Cancer Tissue

作　　者：万敬员[1] 叶笃筠[1] 吴萍[1] 黄云峰[1] 孙伟[1] 应红刚[1] 盘茜[1] 袁萍[1] 张代娟[1] 张力[1] 

机构地区：[1]华中科技大学同济医学院基础医学院病理生理学系,武汉430030

出　　处：《华中科技大学学报（医学版）》2003年第5期467-470,共4页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目 (No .396 70 887;No .30 0 70 92 9)

摘　　要：目的　探讨人结肠癌组织中是否存在环加氧酶 2 (COX 2 )选择性剪接异构体的表达及其可能意义。方法针对人COX 2DNA第 7、 8外显子序列 ,设计一对全新引物 ,并采用RT PCR技术 ,从结肠癌组织中分离COX 2mRNA ,然后对电泳条带进行克隆、测序和分析。结果　正常结肠组织和结肠癌组织均发现COX 2目的条带 ( 2 5 2bp) ,但结肠癌组织则出现了一条新的电泳带 ( 5 34bp) ,经克隆和测序证实 ,该条带除目的条带的COX 2DNA的第7、 8外显子序列外 ,还包含第 7、 8外显子间的内含子序列 ,即保留的内含子 ( 2 82bp)。根据阅读框推测 ,在保留的内含子第 4 8～ 5 0碱基出现终止密码。结论　首次发现结肠癌组织中存在COX 2基因的选择性剪接异构体 (Genbankaccessionnumber:BU4 936 0 2 ,AY15 12 980 ) ,其所编码蛋白可能较已报道的COX 2酶蛋白小 ,且蛋白质的C末端缺少阿斯匹林作用位点。Objective To explore the expression of cyclooxygenase 2 (COX 2) alternative splice variant in human colon cancer tissue and its possible significance. Methods According to the sequence of exons 7, 8 of human COX 2 gene, the special primers were designed and synthesized. Reverse transcription polymerase chain reaction technique was used to detect the expression of COX 2 in human colon cancer tissues. PCR products were cloned, sequenced and analyzed by using BLAST and some bio informatics methods. Results The electrophoresis of the RT PCR product showed one short band of reported COX 2 comprising of 252 bp that contained the exons 7 and 8 in human normal colon tissue and colon cancer tissue, but one long new band comprising of 534 bp was detected in human colon cancer tissue. Sequence analysis showed that on 282 bp intron sequence was inserted between exons 7 and 8. Analysis of the predicted amino acid sequence indicated that an in frame stop codon was contained in the 48-50 bp of retention intron.Conclusion A new COX 2 splice variant that may, at best, code for a truncated COX 2 protein has been identified in human colon cancer tissue. Its biological significance remains to be further investigated.

关 键 词：人结肠癌组织 环加氧酶-2 mRNA剪接异构体 分离 克隆 

分 类 号：R735.35[医药卫生—肿瘤]