玉米抗丝黑穗病候选基因cDNA全长克隆及功能分析

玉米丝黑穗病是由丝轴黑粉菌引发的一种绝产性病害，其抗性受多个数量性状位点控制，在染色体2.09区域已检测到主效抗性位点HSQ2.09，并开发出紧密连锁标记。本项目前期通过生物信息学手段预测出该位点的抗性候选基因，并创造了以高感丝黑穗病玉米自交系黄早四为遗传背景，齐319为供体含主效抗性基因HSQ2.09的近等基因系L282。本研究拟以抗病近等基因系与黄早四为材料，采用丝轴黑粉菌混合交配型菌液针刺茎基部接种技术，通过Real-time PCR技术研究抗玉米丝黑穗病基因HSQ2.09的表达模式；结合RACE技术克隆抗病候选基因全长cDNA序列，对其进行生物信息学分析；在通过体外抑菌实验分析抗病性基础上，构建过量表达载体，转化高感玉米自交系黄早四，验证基因功能，进而阐明玉米抗丝黑穗病基因作用的分子机理。本项目研究结果有助于丰富玉米抗病遗传基础。

Head smut caused by Sporisorium reilianum f. sp. zeae (Kühn) is an important disease in maize. A number of studies evidenced that the behavior of head smut resistance in maize belonged to quantitative trait controlled by many loci. A major QTL HSQ2.09 has been mapped on maize chromosome 2 (bin 2.09), and several linked markers have been developed in this region. Currently, we have developed the near-isogenic line (NIL) carrying HSQ2.09 by backcrossing combined with MAS from donor line Qi319 into the Huangzao4 genetic background. In this project,the expression pattern of resistance candicate genes will be studied using Real-time PCR technique and the NIL and Huangzao4 incoculated by needle puncturing method. Furthermore, the full-length cDNA encoding head smut resistance candidate gene will be cloned via RACE, and structural characters wil be analysed using a bioinformatic. The fuction of the resistance gene will be validated by using prokaryotic expression in vivo and transgenic technique. The purpose of this project is to clone the gene, and reveal the disease-resistant mechanism of head smut in mazie.