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该研究以泸定百合(Lilium sargentiae Wilson)为材料,构建其组培苗经百合尖孢镰刀菌侵染后的叶片SSH文库,从中筛选镰刀菌枯萎病抗病相关基因。从正向SSH文库中随机挑取300个单克隆测序后得到280条ESTs,进行功能比对分析后,除去未知功能、沉冗蛋白以及无同源序列,得到有功能的ESTs共168条,其功能涉及信号传导、蛋白质合成与代谢、抗病与防御、物质与能量代谢、转录相关等多种途径,其中有31条ESTs与抗病防御相关。从抗病防御相关基因中选取8条ESTs:过氧化氢酶、ATP结合盒转运蛋白(ABC transporter)、Kunitz型胰蛋白酶抑制剂4(Kunitz trypsin inhibitor 4)、丝氨酸乙醛酸氨基转移酶、多聚泛素、脂氧合酶I(Lipoxygenase I)、丝氨酸/苏安酸蛋白激酶(Serine/Threonine-protein kinase)、抗坏血酸过氧化物酶(Arabidopsis thaliana),通过RTPCR对其表达情况进行分析,发现经镰刀菌诱导后均为上调表达,推测它们可能参与了泸定百合镰刀菌枯萎病的抗病反应途径。
In this study, Lilium sargentiae Wilson was used as the material to construct the SSH library of the tissue culture seedlings infected with Fusarium oxysporum fusarium so as to screen out the Fusarium wilt disease-related genes. A total of 280 ESTs were obtained from 300 random clones sequenced from the forward SSH library. After functional alignment analysis, unknown ESTs, divergent proteins and no homologous sequences were removed and 168 ESTs were obtained. The functions involved in signal transduction, protein synthesis and metabolism, disease resistance and defense, material and energy metabolism, transcriptional correlation and many other ways, of which 31 ESTs are associated with disease resistance and defense. Eight ESTs were selected from the genes related to disease resistance and defense: catalase, ABC transporter, Kunitz trypsin inhibitor 4, serine glyoxylate aminotransferase, Polyunsulin, Lipoxygenase I, Serine / Threonine-protein kinase and Arabidopsis thaliana were analyzed by RTPCR and the results showed that After being induced by Fusarium, all of them were up-regulated, suggesting that they may be involved in the disease-resistant pathway of Fusarium et al.