正响应盐胁迫的甘蔗6-磷酸葡萄糖脱氢酶基因的克隆

Cloning and characterization of sugarcane 6-phosphogluconate dehydrogenase gene,positive response to salt stress

从甘蔗中克隆获得1个6PGDH基因的cDNA序列,记为Sc6PGDH(登录号:KF921299).该基因序列含有1个1467 bp的完整开放阅读框,推导编码488个氨基酸残基.生物信息学分析可知,Sc6PGDH基因编码的蛋白分子质量为53.6561 ku,为酸性稳定的亲水蛋白,含有信号肽,可能定位于内质网膜上,具有该类蛋白典型的保守结构域6PGD,且与分离自谷子、玉米和水稻的6-磷酸葡萄糖脱氢酶的氨基酸序列同源性分别高达96.11%、95.70%和93.24%,表明该基因在进化过程中比较保守.原核生物生长试验结果显示,甘蔗Sc6PGDH基因的表达蛋白可以显著增强大肠杆菌Rosetta菌株的耐盐性,推测该基因可能参与了生物体对盐胁迫的应答过程.

The16-phosphogheonate dehydmgenase （6PGDH）, a key rate-limiting enzyme of the pentose phosphate pathway in plant, plays a vital role in plant growth and response to adversity stresses. One 6-phosphog, luconate dehydrogenase gene, named as Sc6PCDH （ GenBank Accession No. KF921299）, was isolated from Saccharum complex hybrid using the methods of 6ombination of in silico cloning and RT-PCR technology. The cDNA sequence of Sc6PGDH contained an open reading frame with length of 1467 bp, encoding 488 amino acid residues deduced and the size of Sc6PGDH protein being 53.6561 ku. According to the analysis of bioinfor- matics, the sugarcane Sc6PGDH was an acid stable hydropliilic protein with signal peptides, probably locating on the endoplasmic reticulum membrane. The Sc6PGDH contained two typic conserved domains of 6-phosphogluconate dehydrogenase and shared high homologies with the 6-phosphogluconate dehydrogenases isolated from Ory~ sativa Japonica Group （NP001056586）, Setar/a/ta/ica （XP004971196） and Zea mays （DAA55929）, which was 93.24%, 96.11% and 95.70%, respectively, indicating the 6PGDH gene was relatively conservative in evolution process. The spot assay revealed the Sc6PCDH expressed in Eseherichia coli （ strain Rosetta） could significantly increase the salt tolerance, suggesting the Sc6PGDH gene involved in the positive response to the salt stress.