大豆热激蛋白基因HSP17.4的耐热功能鉴定

Identification of Heat Tolerance of Small Heat Shock Protein Gene HSP17.4 from Soybean

为研究小热激蛋白基因HSP17. 4在大豆热生物胁迫耐受过程中的作用,利用农杆菌介导法,将植物过表达载体p CPB-HSP17. 4和RNA干扰表达载体p CPB-HSP17. 4-RNAi导入受体大豆JN18中。经PCR检测,获得T0代阳性植株13株,T1代阳性植株26株,T2代阳性植株39株。T1、T2代转基因植株Southern blotting结果显示,目标基因以单拷贝形式整合到基因组中。荧光定量PCR结果显示,HSP17. 4基因在转化植株的叶、茎中均有表达。在42℃高温胁迫下,与未转化植株相比,T1、T2转过表达植株叶片中相对表达量分别为未转化植株的813%和793%,转RNAi表达植株叶片中相对表达量分别为未转化植株的49. 77%和52. 81%。耐高温鉴定结果表明,转过表达HSP17. 4基因提高了植株的耐高温能力。

To study the function of a small heat shock protein HSP17. 4 in thermotolerance of soybean（ JN18）,HSP17. 4 gene was cloned from the JN18. An overexpression vector pCPB-HSP17. 4 and an RNA expression vector pCPB-HSP17. 4-RNAi were constructed. Detection of PCR showed that we obtained thirteen transformed plants in T0 generation,twenty-six transformed plants in T1 generation and thirty-nine transformed plants in T2 generation of JN18. Southern blotting of T1 and T2 generations indicated that target gene was integrated into the genome of soybean with single copy.The quantitative real-time PCR showed that the HSP17. 4 gene was expressed in the leaves and stems of transformed plants. Under high temperature stress at 42,compared with the non-transformed plants,the relative expression of T1 and T2 transformed plants in pCPB-HSP17. 4 was 813% and793%,respectively. The relative expression of T1 and T2 transformed plants in pCPB-HSP17. 4-RNAi was 49. 77% and 52. 81% that of non-transformed plants in leaves,respectively. The results showed that the overexpression of HSP17. 4 gene could improve the high temperature resistance of soybean.