中华大蟾蜍mcl-1基因克隆及其重组蛋白在原核体系中的诱导表达  

Bufo gargarizans mcl-1 cloning and its prokaryotic recombinant protein expression

作　　者：胡巧玲[1] 张姝芳[2] 杨仙玉[1] 余美华[1] 诸葛慧[2] 

机构地区：[1]浙江农林大学动物科技学院,浙江临安311300 [2]浙江农林大学亚热带森林培育国家重点实验室培育基地,浙江临安311300

出　　处：《药学学报》2013年第10期1624-1628,共5页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(31071181)

摘　　要：以蟾蜍（Bufo）为基原的常用中药材有蟾酥、蟾衣、蟾皮、干蟾等，具有强心、麻醉、解毒、止痛、抗菌、增强免疫力及抗肿瘤等药理作用，按溶解性可将其成分划分为脂溶性甾族类和水溶性吲哚生物碱类。体内外实验已证明由蟾皮制成的水溶性注射液——华蟾素具有良好的抗肿瘤效果。MCL-1 is encoded by myeloid cell leukemia-1 gene （mcl-1）, which is one of the anti-apoptotic members of bcl-2 cell apoptotic gene superfamily. ChanSu is made of dorsal secretions of several Bufo species, commonly used in the prescriptions of traditional Chinese medicine for treating many diseases including cancer. To clarify if mcl-1 is expressed in the dorsal skin of B. gargarizans, the PCR （polymerase chain reaction） was performed with its dorsal skin first strand cDNA as the template and a pair of specific primers of mcl-1, and PCR products were cloned into the pGM-T vector. DNA sequencing indicated that the ORF length was 639 bp encoding 212 amino acid residues, and the homology of 44%-95% with the MCL-1 of several other animals. For the further studies on MCL-1 biological functions during the oncogenesis and preparation of its antibody, the prokaryotic expression construct of pET-28b-mcl-1 was prepared which was confirmed by DNA sequencing, and its recombinant protein expression （0.02% wet weight） in E. coli BL21 （DE3） strain was confirmed by SDS-PAGE and Western blotting.

关 键 词：髓细胞白血病基因-1 中华大蟾蜍 原核表达 重组蛋白 

分 类 号：R915[医药卫生—微生物与生化药学]