玉米自交系齐319抗灰斑病主效位点qGLS1.02的精细定位

发布时间：2018-01-08 11:36

  本文关键词：玉米自交系齐319抗灰斑病主效位点qGLS1.02的精细定位　出处：《沈阳农业大学》2017年硕士论文　论文类型：学位论文

  更多相关文章： 玉米 灰斑病 染色体片段置换系 精细定位 生物信息学

【摘要】：玉米灰斑病(Gray Leaf Spot,GLS)是一种在全世界广泛分布的玉米真菌性病害,主要由玉蜀黍尾孢(Cercospora zeae-maydis和玉米尾孢(Cercospora zeina)引起,给玉米生产造成了极大危害。鉴定并利用抗病种质资源改良我国骨干自交系以培育抗病品种是防治灰斑病发生的有效途径。本课题组前期利用以抗病自交系齐319和感病自交系掖478构建的包含300个家系的重组自交系(RIL)群体,结合由199个SSR标记构建的遗传图谱,根据2年灰斑病人工接种鉴定,在第1染色体binl.02位置发掘出了一个玉米抗灰斑病(玉蜀黍尾孢)主效QTL,qGLS1.02,两端标记为umc2614和bnlgl803,物理距离长5.73Mb。本试验于2015年以齐319为供体亲本、掖478为轮回亲本构建的19个BC5F3世代共9种不同基因型的染色体片段置换系为试验材料,采用人工接种对其衍生的19个BC5F4世代群体进行灰斑病抗性鉴定,通过T检验验证qGLS1.02初定位区间的抗病性,结果显示来源于齐319的qGLSl.02片段能显著提高掖478对灰斑病的抗性。从2015年的15SY15群体中筛选出的含有qGLS1.02全长杂合片段的BC5F4单株进行自交,将BC5F5世代单株用于重组体筛选,利用加密的分子标记共筛选出了 10种重组体,进一步于2016年将重组体回交衍生的10个BC5F6世代群体进行抗病鉴定,通过群体抗病性分类、卡方检验及T检验等策略将qGLS1.02区间缩小到了 363.61Kb范围内。参照B73测序结果V2版本,根据网站Maize GDB分析结果,在精细定位区间内存在5个基因,进一步对该5个基因进行生物信息学分析,进行候选基因的预测。5个基因中只有KHB5在叶部表达量较高,同时具有明确、一致的功能预测,将其作为预测的候选基因。利用候选基因KHB5临近标记SYLY_80对201份玉米自交系进行基因分型,结合灰斑病抗性的鉴定结果,通过T检验以验证该标记对玉米灰斑病抗性选择的有效性,结果显示与齐319基因型相同的自交系整体抗病性显著优于其它自交系,说明该标记能够对玉米灰斑病抗性进行有效选择。通过对qGLS1.02主效基因的精细定位、生物信息学预测及分子标记的有效性验证为揭示玉米抗灰斑病菌的机理、分子标记辅助育种的应用提供了理论基础。
[Abstract]:Gray Leaf spot (GLS) is a widely distributed fungal disease in maize all over the world. It is mainly caused by Cercospora zeae-maydis and Cercospora zeina. It is an effective way to control gray spot disease by identifying and using resistant germplasm resources to improve Chinese backbone inbred lines to cultivate disease-resistant varieties. A recombination inbred line consisting of 300 families was constructed from Ye478, a susceptible inbred line. The Ril group. Based on the genetic map constructed by 199 SSR markers, two years' work inoculation of gray spot patients was carried out. A dominant QTL QGLS1.02 was identified at the binl.02 site on chromosome 1 in maize resistant to gray spot disease (Cercospora zebra). The two ends were labeled as umc2614 and bnlgl803, and the physical distance was 5.73 Mb. in this experiment, Qi319 was used as donor parent in 2015. The chromosome fragment replacement lines of 9 different genotypes in 19 BC5F3 generations constructed by Ye 478 as recurrent parents were used as experimental materials. The resistance to gray spot disease was identified by artificial inoculation in 19 BC5F4 populations derived from it, and the disease resistance of qGLS1.02 initial locational interval was verified by T test. The results showed that the qGLSl.02 fragment derived from Qi319 could significantly increase the resistance of Y478 to gray spot disease. A total of qGLS1.02 was selected from 15SY15 population in 2015. The single BC5F4 plant with long heterozygous fragment was self-pollinated. BC5F5 generation single strain was used to screen the recombinant, and 10 kinds of recombinant were screened by using the encrypted molecular marker. In 2016, 10 BC5F6 generation populations derived from the recombinant were identified and classified by population disease resistance. Chi-square test and T test reduced the qGLS1.02 range to 363.61Kb, referring to the V2 version of B73 sequencing results. According to the results of Maize GDB analysis on the website, there were 5 genes in the fine mapping region, and the five genes were further analyzed by bioinformatics. Among the 5 genes, only KHB5 expression was higher in the leaves, and had a clear and consistent function prediction. The candidate gene was used as the candidate gene for prediction. 201 maize inbred lines were genotyped by using the candidate gene KHB5 adjacent marker SYLY_80, and the results of identification of resistance to gray spot disease were combined. T-test was used to verify the effectiveness of the marker in the selection of resistance to gray spot disease in maize. The results showed that the overall resistance of inbred lines with the same genotype as Qi319 was significantly better than that of other inbred lines. The results showed that the marker could be used to select the resistance to gray spot of maize effectively. The fine mapping of the main gene of qGLS1.02 was carried out. Bioinformatics prediction and validity verification of molecular markers provide a theoretical basis for revealing the mechanism of resistance to gray spot fungi in maize and the application of molecular marker assisted breeding.
【学位授予单位】：沈阳农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S435.131.4
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本文编号：1396963