割手密是栽培种甘蔗最重要的近缘物种之一,具有较强的抗逆性。本研究以高粱过氧化氢酶基因(NCBI登录号为XM002437586.1)c DNA序列为探针,对甘蔗EST数据库进行同源检索、筛选和拼接,后经基因组PCR、分子克隆、序列分析验证成功分离了2个割手密过氧化氢酶基因c DNA序列(Ss CAT-1c和Ss CAT-1d,Gene Bank登录号为KF864229和KF864230)。2条序列全长均为1 532 bp,包含10 bp的5'非翻译区(UTR)和43 bp的3'UTR,以及一个1 479 bp的开放读码框,编码492个氨基酸,此蛋白序列具有过氧化氢酶保守结构域,属于CAT家族。2条序列之间相似性为99.2%,存在12个单核苷酸多态性(SNP)位点,蛋白相似性99.0%,存在5个氨基酸变异位点。将推测的Ss CAT-1c和Ss CAT-1d蛋白序列与高粱、水稻、玉米等物种进行同源进化分析,均表现出极高的保守性。研究结果能对今后深入了解甘蔗及其近缘材料CAT基因家族的信息提供帮助,从而为进一步通过生物技术改良甘蔗品种抗逆性提供理论依据。
Saccharum spontaneum L. is one of the most important wild sugarcane germplasm having strong resistance. In this research, two novel full-length cDNA sequences of catalase (CAT) gene (SsCA T-1c and SsCA T-1d, GenBank accession number KF864229-30) were cloned by in silico cloning and experimentally validated. The cDNA length of SsCA T-1c and SsCA T-1d all was 1 532 bp, containing 10 bp of 5'' untranslated region (UTR) and 43 bp of the 3''UTR, and an open reading frame of 1 479 bp encoding a polypeptide of 492 amino acid. The protein encoded by SsCA T-1c and SsCA T-1d contains a catalase family conserved domain. The high homology of cDNA (99.2%) and protein (99.0%) were found between SsCA T-1c and SsCA T-1d. There are 12 single nucleotide poly-morphisms (SNP) sites for cDNA, and 5 amino acid mutation sites for protein between SsCA T-1c and SsCA T-1d. The putative proteins encoded by SsCAT-1c and SsCAT-1d have high homology with sorghum, rice, corn and other species.