【作者】 刘晓维；

【导师】 徐敬友；

【作者基本信息】 扬州大学 ， 植物病理学， 2010， 硕士

【摘要】 植物多聚半乳糖醛酸酶抑制蛋白(PGIP)是一种能够特异性识别和结合真菌内切多聚半乳糖醛酸酶(PG)并抑制其活性的蛋白,其富含亮氨酸重复(LRR),在植物抗真菌过程中发挥重要作用。本研究以水稻品种Lemont、Jasmine85和YSBR1为材料,克隆水稻PGIP基因(Ospgip1),并进行原核表达,同时明确OsPGIP1对水稻纹枯病菌PG的抑制活性。另外,通过荧光定量PCR,分析Ospgip1在水稻不同抗、感纹枯病品种中的时空表达,以及黄化处理和病菌侵染对Ospgip1的诱导表达,从而探明PGIP表达量与品种抗病性的关系。具体结果如下:1、通过PCR技术,克隆获得Ospgip1基因。该基因全长为1 184 bp,具有930 bp的完整开放阅读框(ORF),终止密码子为TAA,无内含子。生物信息学分析表明,OsPGIP1由309个氨基酸组成,含9个LRR重复,1个疏水性N端信号肽;二级结构LRR区含有α-螺旋和β-折叠各9个,分别位于LRR首尾,此外,不规则盘绕(卷曲)占有较高比例(53%);三级结构倾向于产生纤维状构象。2、RT-PCR研究表明,Ospgip1在水稻抗、感纹枯病品种中均能表达,但在不同生育期、不同部位是否表达则有差异。根据Real Time PCR测定,水稻抗病品种YSBR1和Jasmine85中Ospgip1的表达量要明显高于感病品种Lemont,表明Ospgip1表达与品种抗病性有一定关系。3、Real Time PCR研究显示,稻苗黄化处理后,无论是抗病品种还是感病品种的Ospgip1表达均显著提高。但是,纹枯病菌侵染使得抗病品种Ospgip1表达量大大增加,而对感病品种影响不大。更多还原

【Abstract】 Polygalacturonase inhibiting proteins (PGIPs) are anti-fungal proteins with leucine-rich repeat (LRR), which could inhibit the catalytic activity of fungal endo-PG.In this study, gene Ospgip1 encoding OsPGIP1 in rice plants was cloned by PCR. The sequence and structure of OsPGIP1 was infered by biological information analysis and its inhibition to the activity of Rhictonia solani PG was surveyed. Furthermore, Ospgip1 expression in E. coli, its time and spatial expression in rice plants and its expression induced by the yellowing treatment and the pathogen inoculation were conducted. The detailed results are as follows:1. Ospgip1 was a sequence of 1 184 bp, which contained a complete ORF of 930bp in length and encoded a protein of 309 amino acids with hydrophobic N-terminal signal peptide and 9 LRR, called OsPGIP1. The secondary structure of OsPGIP1’s LRR had 9α-helixes and 9β-sheets that sat at the front or end of LRR, and more irregular coils (53%). At last, the protein formed one fibriform structure.2. According to RT-PCR, OsPGIP1 could be constritutively expressed in rice varieties resistant and susceptible to R. solani, but there was some difference in the expression in different growth stages or organs. Real Time PCR indicated that OsPGIP1 was expressed in rice resistant varieties (YSBR1 and Jasmine85) significantly higher than in the susceptible one (Lemont), which showed that there was a postive relationship of OsPGIP1 expression with the resistance of rice varieties.3. OsPGIP1 expression induced by the yellowing treatment and the pathogen inoculation was studied by Real Time PCR. The results showed that OsPGIP1 expressed in rice seedlings was recreased after the yellowing treatment. But, in the infection of R. solani, OsPGIP1 exprossion in the resistant varieties was significantly stronger than that in the control, and no change occurred in the exprossion in the susceptible one.更多还原