麻疯树水通道蛋白新基因JcPIP干旱胁迫下的功能分析

采用RT-PCR和RACE技术,从大戟科(Euphorbiaceae)耐旱植物麻疯树(Jarropha curcas)cDNA中克隆得到了一个麻疯树质膜内膜蛋白(PIP)新基因,命名为JcPIP.聚类分析表明,麻疯树PIP蛋白与蓖麻、葡萄和菠菜的PIP蛋白在进化上有最近的亲缘关系.将JcPIP基因在非洲爪蟾卵母细胞(xenopus oocytes)中异源表达发现细胞膨胀率扩大了10倍,表明JePIP基因编码的是一个水通道蛋白.合成亲水性、抗原性好的JcPIP保守序列多肽,制备并纯化JcPIP多克隆抗体.Western-bbt检测显示JcPIP蛋白富集在29 kDa区段,且在麻疯树各组织中均有大量表达.PEG-6000干旱胁迫下麻疯树叶片JcPIP蛋白丰度增加,复水后丰度开始下降,表明JcPlP与麻疯树的耐旱性相关.

Cloning of a New Aquaporin Gene(JcPIP) from Jatropha Curcas and Analysis of Its Function under Drought Stress

cDNA of Jatropha curcas was amplified by PCR with degenerated primers, a new plasma membrane intrinsic protein gene (JcPIP) was cloned. Phylogenic analysis indicated that PIPs of Jatropha curcas was closer relationship to those in Ricinus communis, Vitis vinifera and Spinacia oleracea. The cRNA of JcPIP was hetero expression in xenopus oocytes as a water channel and increased swelling rate by 10 times. A 13 amino acid sequence peptide was selected from the hydropilic conserved peptides. Then a high titer polycolonal antibody was obtained by immunizing rabbits with the peptide. ELISA analysis showed the titer of rabbit serum against anti-peptide of JcPIP was up to 10-4. The location of JcPIP in Jatropha curcas was analysised with western-blot. JcPIP was major in 29 kDa band and located in all plant tissues. Treated with PEG-6000, the amount of JcPIP incresed, however, decreased under the conditon of rewater. It suggested that JcPIP related with drought tolerance of Jatropha curcas.