日本蟾蜍皮肤胸腺素α原cDNA的克隆及序列分析

Cloning and sequence analysis of prothymosin-α cDNA of Bufo japonicus formosus

为研究蟾酥、蟾衣、蟾皮中多肽类有效成分,通过菌落聚合酶链式反应(polymerase chain reaction,PCR)对日本蟾蜍Bufo japonicus formosus皮肤cDNA质粒文库进行了筛选,获得胸腺素α原(prothymosin-α,ProTα)全长cD-NA序列,并对它们进行了生物信息学分析。日本蟾蜍ProTαcDNA全长为1 480 bp,包括339 bp的完整开放阅读框(open reading frame,ORF),5′端125 bp及3′端1 016 bp的非翻译区(untranslated region,UTR)。根据cDNA序列推导的日本蟾蜍ProTα由112个氨基酸残基组成,无信号肽结构。氨基酸序列同源性分析显示,日本蟾蜍ProTα与食用蛙Rana esculenta同源性为82%,与其他11种动物的同源性则介于54%～73%。ProTα具有显著的抗肿瘤作用。分析日本蟾蜍ProTαcDNA序列,可为研究其生物学功能和药物研发提供实验依据。

To study the polypeptide effective components included in the skin of Bufo japonicus formosus （Japanese toad） and their secretions, the full length cDNAs were screened from the plasmid cDNA library of adult Japanese toad skin using colony polymerase chain reaction with a pair of primers, SP6 （the upstream primer of the vector） and poly（T） （a self-designed primer recognizing the area connecting polyA tail of cDNA and the vector）. Results showed that a prothymosin-α （ProTα） cDNA was obtained from 1 344 colonies checked, whose transcript was 1 480 bp in length consisting of 125 bp in the 5 prime untranslated region （5＇ UTR） and 1 016 bp 3＇ UTR with a complete open reading frame （ORF） of 339 bp encoding a polypeptide of 112 amino acid residues. The homologous analysis indicated a similarity between B. japonicus formosus and Rana esculenta of up to 82%, but for other species it was 54%-73%. Previous studies have shown significant anti-tumor effects of ProTα; therefore, this study could contribute to further studies on ProTct. [Ch, 4 fig. 17 ref.]