水稻OsGPDH1的克隆与功能鉴定

Cloning and Functional Identification of OsGPDH1 in Rice

为了解水稻甘油-3-磷酸脱氢酶基因在水稻抗逆性中的作用,本研究从水稻品种日本晴中克隆了1个编码甘油-3-磷酸脱氢酶的基因,命名为OsGPDH1,该基因编码的蛋白酶具有NAD(P)+结合域和脱氢酶域,且这2个结构域在植物中高度保守。构建过表达OsGPDH1基因载体转化水稻得到转基因植株,RT-qPCR分析表明,OsGPDH1基因在水稻孕穗期的叶、幼穗、茎、节、叶鞘中均有表达,说明该基因参与了水稻的生长发育过程。OsGPDH1基因也受到PEG6000、高盐、双氧水等逆境和甲基茉莉酸(mJA)、水杨酸(SA)等激素诱导表达,且诱导12h后,OsGPDH1的表达量达到最高水平,但对脱落酸(ABA)不敏感。盐胁迫下的发芽试验表明,过表达转基因水稻的发芽率高于野生型,说明OsGPDH1基因表达量的提高可增强转基因植株对盐胁迫的耐受性。对过表达OsGPDH1的转基因水稻进行苗期20%PEG6000胁迫处理后,转基因水稻的成活率显著高于野生型,说明OsGPDH1基因过表达可提高水稻苗期抗旱能力。本研究初步证明了OsGPDH1水稻抗盐胁迫和渗透胁迫的重要作用,为培育抗性转基因水稻新品种提供了新的基因资源。

In understand the roles of glycerol-3-phosphate dehydrogenase gene in stress tolerance in rice, a gene encoding glycerol-3-phosphate dehydrogenase, named as OsGPDH1, was cloned from Nipponbare（Oryza sativa L. ssp. Japonica）in this study. OsGPDH1 encodes a protein that has NAD（P）＋ binding domain and dehydrogenase domain. And the two domains are highly conserved in plants. The real-time RT-PCR analysis showed that OsGPDH1 was expressed in flag leaf, spikelet, stem, sheath, node of adult plant, which indicates the gene involved in the process of growth and development of rice. After stress treatments, OsGPDH1 could be induced by PEG6000, salt, hydrogen peroxide, methyl jasmonate （m-JA） as well as salicylic acid （SA）, and its expression often reached the peak after 12h. However, OsGPDH1 was insensitive to ABA. The overexpressing gene vector was constructed and transferred into rice, and transgenic plants were obtained. The germination rate of overexpressional plants was higher than the wild-type rice under salt treatment, indicating that enhancing OsGPDH1 expression can improve tolerance to salt stress. The survival of over-expression transgenic plants was significantly higher than of the wild-type plants under 20% PEG6000 treatment at seedling stage, indicating that enhancing OsGPDH1 expression can improve the drought-resistance at the seedling stage. This study demonstrated that OsGPDH1 played an important role in salt and osmotic stress, and could be a new transgenic genetic resource for stress resistant breeding in rice.