Lenzites gibbosa锰过氧化物酶1基因启动子序列的克隆

Cloning of Lg-mnp1 Gene Promoter from Basidiomycete Lenzites gibbosa

根据已克隆到的Lenzites gibbosa锰过氧化物酶1cDNA全长基因Lg-mnp1,在5'端设计了3个巢式特异性引物,利用染色体步移技术克隆得到了其上游1 568 bp的启动子序列。采用Signal Scan进行了启动子序列分析。结果表明,在起始密码子上游-92～-43 bp区域为Lg-mnp1启动子的基础启动子区;在-52 bp处有一个转录起始位点,-83 bp处有1个TATAAA-box,-119、-196 bp有2个反向CCAAT-box(ATTGG)。启动子区也存在多个顺式作用元件,包括转录因子SP-1(GGGCGG)、转录因子AP-2(CCCMNSSS)、热激元件HSE(NTTCNNGAAN)及6个推定的金属响应元件MRE(TGCRCNC)等。

Three nested sequence-specific primers were designed according to the full length sequence of Lg-mnpl eDNA cloned by ourselves. A 1 568-bp upstream promoter sequence was cloned by the chromosome walking method and analyzed using the online software Signal Scan. Results showed that the core promoter sequence was predicted at from -92 to -43 bp, the transcription start located at -52 bp, the TATAAA element located at -83 bp, and two inverted GCAAT elements （ATF- GG） located at -119 bp and -196 bp upstream of the translation initiation codon of Lg-mnpl. There were a number of other cis-acting elements, including consensus SP-1 transcription factor recognition site （ GGGCGG）, the consensus for the binding of transcription factor AP-2 （ CCCMNSSS）, one HSE matching the consensus 5＇-NrITCNNGAAN-3＇, and 6 putative MREs identical to the consensus 5＇-TGCRCNC-3＇.