摘要
大豆易遭受干旱、盐、低温等非生物胁迫,严重导致产量下降。利用分子生物学技术提高作物抗性是作物育种的有效途径。锌指蛋白是植物中常见的重要转录因子,能够调控多种胁迫诱导基因的表达,有效提高综合抗性。在这项研究中,利用RT-PCR方法克隆大豆(Glycine max L.)GmWRKY35基因。其cDNA为1 500 bp,编码一个499个氨基酸的蛋白质,预测其分子量为54.89 kD,等电点(p I)为6.74。亚细胞定位分析表明,GmWRKY35定位于细胞核。实时荧光定量PCR分析显示,GmWRKY35转录能被干旱诱导。把GmWRKY35基因通过农杆菌介导转化烟草(Nicotiana tabacum L.)中。在干旱胁迫下,与野生型烟草相比,转基因烟草植株表现出主根较长,叶子萎焉少,POD和SOD活性较高,MDA含量和电解质渗漏率较少。主要功能验证表明,GmWRKY35基因在烟草中表达增强了干旱胁迫耐受性。这项研究表明大豆RING-H2型锌指蛋白在植物逆境中有重要作用,同时也为大豆抗性育种提供一个优良的候选基因。
Soybean, like other crops, suffers from drought, salt, low-temperature and other abiotic stresses, seriously resul- ting in yield decline. It is an effective way for crop breeding to improve resistance via molecular biology techniques. Zinc fin- ger protein, an important transcription factor commonly found in plant, can regulate the expression of multiple stress-inducible genes and enhance comprehensive resistances effectively. In this study, a RING-H2 zinc finger protein gene, designated as GmWRKY35, was isolated from Glycine max L. by RT-PCR. Its cDNA was 1 500 bp and encoded a putative protein of 499 amino acids with a predicted molecular mass of 54. 89 kD and an isoelectric point （PI） of 6. 74. A typical RING-H2 finger domain was found at the C-terminal region of GmWRKY35 protein. Subcellular localization analysis showed that the GmWRKY35 was expressed in nuclear. Real-time PCR showed that the transcript of GmWRKY35 was strongly induced by drought, salinity and cold to some extent. The GmWRKY35 gene was transformed into tobacco cuhivar Wisconsin 38 （W38） by Agrobacterium mediated under the control of the CaMV 35S promoter. Under drought stresses, transgenic tobacco lines car- rying GmWRKY35 gene developed the strong primary root, smaller yellow leaves, higher POD and SOD activity, less MDA content and the percentage of electrolyte leakage, and performed higher tolerance to these stresses than the wide-type tobac- cos. The primary function verification showed that overexpression of GmWRKY35 gene in tobaccos enhanced their tolerance to drought stresses. This study shows that RING-H2 zinc finger proteins of soybean play an important role in plant stress resist- ance and provide an excellent candidate for resistance breeding in soybean.
出处
《大豆科学》
CAS
CSCD
北大核心
2017年第5期685-691,共7页
Soybean Science
基金
河南省科技发展计划(112300410042)
