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受青枯菌诱导的花生根酵母双杂交文库构建和AhRRS5互作蛋白的筛选 被引量:9
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作者 陈玉婷 刘露 +6 位作者 楚盼盼 魏嘉贤 钱慧娜 陈华 蔡铁城 庄伟建 张冲 《作物学报》 CAS CSCD 北大核心 2021年第11期2134-2146,共13页
前期研究报道超表达花生AhRRS5基因能够显著提高烟草抗青枯病水平,为进一步探究NBS-LRR类抗病蛋白AhRRS5在花生应答青枯菌胁迫的信号通路,本研究在构建花生受青枯菌诱导的根部组织均一化三框文库的基础上,通过酵母双杂交技术筛选AhRRS5... 前期研究报道超表达花生AhRRS5基因能够显著提高烟草抗青枯病水平,为进一步探究NBS-LRR类抗病蛋白AhRRS5在花生应答青枯菌胁迫的信号通路,本研究在构建花生受青枯菌诱导的根部组织均一化三框文库的基础上,通过酵母双杂交技术筛选AhRRS5的互作蛋白。通过改良的CTAB法提取青枯菌诱导后不同时间点的花生根部组织样品总RNA,分离纯化mRNA并合成双链cDNA,并基于同源重组方法分别构建酵母双杂交初级和次级文库。构建的酵母双杂交次级cDNA文库库容为1.44×10^(7) cfu mL^(-1),重组率为100%,插入片段大小在1000 bp以上。通过酶切连接法构建pGBKT7-AhRRS5诱饵载体,在酵母细胞中无自激活和毒性活性,与酵母双杂交文库共转酵母Y2H Gold菌株后,经多次筛库和回转验证,最终获得12个候选互作蛋白,这些蛋白涉及到植物生长发育、能量代谢、激素信号转导、胁迫响应等多个方面。通过双分子荧光互补(bimolecular fluorescence complementation,BiFC)验证了AhSBT1.6和AhRRS5的体内互作。转录组数据显示,花生AhSBT1.6基因在不同组织中表达差异显著;实时荧光定量PCR显示,该基因在抗青枯病花生品种中受青枯菌诱导上调表达,推测AhSBT1.6可能参与调控花生青枯病抗性。研究结果为进一步研究NBS-LRR类抗病蛋白AhRRS5和互作蛋白在花生青枯病抗性防御的作用机制奠定了基础。 展开更多
关键词 花生 青枯病 酵母双杂交 互作蛋白 AhSBT1.6 双分子荧光互补
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Phylogenetic Relationships in Genus Arachis Based on SSR and AFLP Markers 被引量:4
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作者 TANG Rong-hua zhuang wei-jian +5 位作者 GAO Guo-qing HELiang-qiong HAN Zhu-qiang SHAN Shi-hua JIANG Jing LI Yang-rui 《Agricultural Sciences in China》 CAS CSCD 2008年第4期405-414,共10页
Fourteen wild species of different sections in the genus Arachis and 24 accessions of the AABB allotetraploid A. hypogaea (cultivated peanut) from several countries which belong to different botanical varieties, wer... Fourteen wild species of different sections in the genus Arachis and 24 accessions of the AABB allotetraploid A. hypogaea (cultivated peanut) from several countries which belong to different botanical varieties, were analyzed by SSR and AFLP marker systems. The assay-units per system needed to distinguish among all the tested accessions were at least five for SSR or two for AFLP. The genetic distance detected by the SSR markers ranged from 0.09 to 0.95, and the mean was 0.73; and the genetic distance detected by the AFLP markers ranged from 0.01 to 0.79 with an average of 0.42. All the tested peanut SSR primer pairs were multilocus ones, and the amplified fragments per SSR marker in each peanut genome ranged from 2 to 15 with the mean of 4.77. The peanut cultivars were closely related to each other, and shared a large numbers of SSR and AFLP fragments. In contrast, the species in the genus Arachis shared few fragments. The results indicated that the cultivated peanut (A. hypogaea L.) varieties could be partitioned into two main groups and four subgroups at the molecular level, and that A. duranensis is one of the wild ancestors of A. hypogaea. The lowest genetic variation was detected between A. cardenasii and A. batizocoi, and the highest was detected between A. pintoi and the species in the section Arachis. The relationships among the botanical varieties in the cultivated peanut (A. hypogaea L.) and among wild species accessions in section Arachis and those in other sections in the genus Arachis were discussed. 展开更多
关键词 ARACHIS SSR AFLP genetic diversity phylogenetic relationship
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Effects of Suppressing OsCRY1a Gene Expression on Rice Agronomic Traits
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作者 LI Yu zhuang wei-jian +5 位作者 WANG Nai-yuan DAI Fei HONG Guo-qin XIE Na-ying LIN Jian-qin QIU Xiu-li 《Rice science》 SCIE 2012年第1期8-13,共6页
Using primers designed according to the published sequence of rice OsCRY1a gene, we obtained part of the gene fragment by PCR and constructed an RNA interference expression vector with it. To down-regulate the express... Using primers designed according to the published sequence of rice OsCRY1a gene, we obtained part of the gene fragment by PCR and constructed an RNA interference expression vector with it. To down-regulate the expression level of the gene or lead to the loss-of-function of the gene, the vector was then introduced into rice via Agrobacterium-mediated transformation. Based on the performance of the transgenic plants, the functions of the gene were analyzed and deduced. The results indicated that suppressing the expression of the gene retarded flowering for 16 d in rice with the plant height and grain length significantly increasing whereas other important agronomic traits observed remained unchanged apparently. 展开更多
关键词 RICE cryptochrome gene OsCRYla RNA interference agronomic trait gene expression
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