CAJ | 학술논문

　　为研究高效产氢菌株成团泛菌(Pantoea agglomerans)BH-18中依赖型磷酸甘油酸变位酶(cofactor-dependent phosphoglycerate mutase, dPGM)与产氢之间的关系,本研究根据GenBank上已登录的肠杆菌中编码 dPGM的基因序列设计一对引物,从细菌基因组 DNA中克隆得到编码 dPGM基因的完整开放阅读框,其长度为753 bp,编码250 aa。采用BLAST对其与NCBI GenBank中的核苷酸序列进行比较分析,结果表明该基因保守性相对较高,与肠杆菌科众多菌株中的 dPGM 基因相似性达100%。采用Bioedit和Mega4软件构建NJ系统进化树,结果表明成团泛菌BH-18的dPGM氨基酸序列与Enterobacter asburiae的dPGM聚为一类,而与成团泛菌属中其他菌株的该蛋白关系较远, dPGM的氨基酸序列在属内不保守。最后,根据已获得的基因序列设计引物,采用RT-PCR的方法分析了成团泛菌BH-18产氢过程中dPGM基因的转录情况,结果表明dPGM基因的转录与产氢呈正相关,依赖型磷酸甘油酸变位酶是产氢过程中的关键酶。
　　위연구고효산경균주성단범균(Pantoea agglomerans)BH-18중의뢰형린산감유산변위매(cofactor-dependent phosphoglycerate mutase, dPGM)여산경지간적관계,본연구근거GenBank상이등록적장간균중편마 dPGM적기인서렬설계일대인물,종세균기인조 DNA중극륭득도편마 dPGM기인적완정개방열독광,기장도위753 bp,편마250 aa。채용BLAST대기여NCBI GenBank중적핵감산서렬진행비교분석,결과표명해기인보수성상대교고,여장간균과음다균주중적 dPGM 기인상사성체100%。채용Bioedit화Mega4연건구건NJ계통진화수,결과표명성단범균BH-18적dPGM안기산서렬여Enterobacter asburiae적dPGM취위일류,이여성단범균속중기타균주적해단백관계교원, dPGM적안기산서렬재속내불보수。최후,근거이획득적기인서렬설계인물,채용RT-PCR적방법분석료성단범균BH-18산경과정중dPGM기인적전록정황,결과표명dPGM기인적전록여산경정정상관,의뢰형린산감유산변위매시산경과정중적관건매。
In order to study the relationship between high hydrogen-production and cofactor-dependent phosphoglycerate mutase (dPGM) in Pantoea agglomerans BH-18, a set of primers were designed based on the dPGM-encoding gene sequences of Enterobacteriaceae which have accession number on GenBank. And the dPGM-encoding gene was obtained by PCR amplification from the bacterial genomic DNA. A complete open read-ing frame of the gene was 753 bp and encoded 250 aa. The results of BLAST analysis in GenBank indicated that the nucleotide sequence of the gene was highly conserved, and the similarity to many strains of Enterobacteriaceae reached 100%. Meanwhile, The NJ phylogenetic tree was constructed by the software of Bioedit and Mega4, and the results indicated that the amino acid sequences of dPGM in BH-18 and Enterobacter asburiae belonged to the same class, whereas it was distant from the protein of the other Pantoea sp. The results suggested the amino acid sequence was not conserved in genus. Then another set of primers were designed according to the dPGM-encoding gene sequence of strain BH-18. The transcription changes of dPGM-encoding gene were analyzed by the method of RT-PCR in the process of hydrogen production. The results demonstrated that the transcription of dPGM gene was positively correlated with hydrogen production, which suggested that dPGM was a key enzyme in hydrogen pro-duction.