摘要：

为探究大豆GmPP2C89基因在植物非生物胁迫响应和适应过程中的功能,利用转录组数据和荧光定量PCR方法检测GmPP2C89基因在NaCl、PEG和甘露醇处理下的表达模式;接着分析GmPP2C89基因启动子上响应非生物胁迫的顺式作用元件,并根据元件的分布克隆不同长度的启动子构建融合GUS载体,获得对应的转基因拟南芥,通过GUS染色分析启动子对NaCl、PEG和甘露醇处理的响应。构建GmPP2C89基因过表达的转基因拟南芥,在正常条件和NaCl处理条件下测定根长、叶片MDA含量和电解质渗透率以及盐胁迫相关基因(SOD、POD、CAT、RD26、RD29A和RD29B)的表达水平。结果显示,NaCl、PEG和甘露醇处理均导致大豆GmPP2C89基因表达水平显著上调;在其启动子区含有较多ABRE、DRE、G-box、MBS、MYB、MYC和TC-rich repeats等参与非生物胁迫响应的顺式作用元件,并且该启动子对NaCl处理具有更强的响应。此外,在盐处理条件下,转基因拟南芥GmPP2C89-OX根长显著大于WT,而MDA含量和电解质渗透率显著低于WT,耐盐性明显增强;抗氧化酶基因(SOD和POD)和ABA途径关键基因RD29B在GmPP2C89-OX中的表达水平显著高于WT。结果表明,大豆GmPP2C89基因受NaCl、PEG和甘露醇诱导表达上调,GmPP2C89过表达可通过激活抗氧化途径和ABA途径增强转基因拟南芥的耐盐性。

关键词: 大豆, GmPP2C89基因, 启动子, 顺式作用元件, 耐盐性

Abstract:

To investigate the function of soybean GmPP2C89 gene in plant abiotic stress response and adaptation. The expression patterns of GmPP2C89 under NaCl,PEG and mannitol treatments were detected by transcriptome data and Real-time quantitative PCR. Then,the cis-acting elements on the promoter of GmPP2C89 in response to abiotic stress were analyzed,and promoters of different lengths were cloned according to the distribution of cis-elements to construct fusion GUS vectors to obtain the corresponding transgenic Arabidopsis. The response of the promoters to NaCl,PEG and mannitol was analyzed by GUS staining. Transgenic Arabidopsis overexpressing the GmPP2C89 was constructed,and the root length,leaf MDA content and electrolytic leakage,and the expression of salt stress-related genes(SOD,POD,CAT,RD26,RD29A,and RD29B)were measured under normal and NaCl treatment conditions. The results showed that NaCl,PEG and mannitol treatments all led to a significant increase in the expression level of soybean GmPP2C89;the promoter region contained many cis-acting elements such as ABRE,DRE,G-box,MBS,MYB,MYC and TC-rich repeats which were involved in abiotic stress response,and this promoter was more responsive to NaCl treatment. In addition,under the salt treatment,the root length of transgenic Arabidopsis GmPP2C89-OX was significantly greater than that of WT,while the MDA content and electrolytic leakage were significantly lower than those of WT,and the salt tolerance was significantly enhanced;the expression of antioxidant enzyme genes(SOD and POD)and ABA pathway key gene RD29B in GmPP2C89-OX was significantly higher than that in WT. These results indicated that soybean GmPP2C89 was induced by NaCl,PEG and mannitol,and GmPP2C89 overexpression could enhance the salt tolerance of transgenic Arabidopsis by activating antioxidant and ABA pathways.

Key words: Soybean, GmPP2C89 gene, Promoter, cis-acting element, Salt tolerance