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作 者 ：袁梦求,丁春邦,陶亮,陈泓翰,陈惠,吴琦

期 刊 ：中草药 2012年 11期 页码：2250-2256

关键词：滇重楼；环阿屯醇合酶；基因克隆；序列分析；开放阅读框(ORF)；

Keywords:Paris polyphylla var. yunnanensis (Franch.) Hand. - Mazz., cycloartenol synthase (CAS), gene cloning, sequence analysis, open reading frame (ORF),

摘 要 ：目的获取滇重楼甾体皂苷合成途径关键酶环阿屯醇合酶基因(PpCAS)的全长cDNA序列，并进行序列分析。方法利用同源克隆和RT-PCR技术获得PpCAS基因保守片段，采用RACE技术获得PpCAS基因的3’及5’末端序列，并采用生物信息学方法进行序列分析。结果 PpCAS基因全长cDNA为2 309 bp，其开放阅读框(ORF)为2 283 bp，可编码760个氨基酸的蛋白质；PpCAS推导的蛋白质相对分子质量为8.69×104，等电点(pI)为6.54；其氨基酸序列与GenBank中其他植物CAS的同源性在60%～83%PpCAS蛋白。结论从滇重楼中首次获得PpCAS基因cDNA全长序列，该基因具有CAS同源基因的典型特征。

Abstract:Objective To clone and analyze the full-length cDNA sequences of cycloartenol synthase gene from Paris polyphyllavar. yunnanensis(PpCAS). MethodsUsing homology cloning and RT-PCR techniques, conserved fragments of PpCAS gene were obtained. RACE technology was usedto obtain 3’ and 5’ end sequences of PpCAS and sequence analysis was done by bioinformatics method. Results The full-length cDNA of PpCAS was 2 309 bp, which contained an open reading frame (ORF) of 2 283 bp. Sequence analysis indicated that PpCAS could encod protein with 760 amino acids, with a predicted relative molecular weight of 8.69 × 104 and an isoelectric point (pI) of 6.54. Sequence analysis and homology modeling suggested that amino acidsequence of PpCAS showed high homology (60%―83%) with CAS of other plants. Conclusion The full-length cDNA sequence of PpCAS is first obtained and it has the typical characteristics of the homologous genes.

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