油菜近等基因系鉴定及BnWRKY16、BnWRKY52功能初探

发布时间：2018-04-04 17:58

  本文选题：近等基因系　切入点：SSR分子标记　出处：《山西大学》2016年硕士论文

【摘要】：油菜是中国主要种植的油料作物,对我国国民经济的发展有着重要的作用,提高油菜的油分含量一直是油菜育种研究的首要任务。WRKY转录因子作为一种调控因子在植物调控网络中占据着重要地位,参与植物的生长发育、抗逆、抗病等一系列的调控过程。我们通过转录组测序和分析得到了两个与含油量有关的WRKY转录因子BnWRKY52、BnWRKY52。本实验利用高代自交姐妹系构建了两个高含油量和中低含油量的近等基因系,并以这两个近等基因系材料为基础材料,初步研究了BnWRKY52、BnWRKY16在油菜种子成熟与萌发过程中的表达情况。具体情况如下:1.以NJ9高油品系为基础材料,利用连续自交的方法构建含油量差异的拟近等基因系。以来源于芸薹属Brassica DB公共数据库的156对SSR引物对9个拟近等基因系株系材料进行扩增,筛选出32对条带清晰、重复性好的引物用于近等性分析。基于32对SSR引物PCR扩增结果,经过遗传和似性系数估算以及聚类分析鉴定近等基因系的近等性,结果显示9个候选株系材料遗传相似性系数均在0.85到0.94之间,其中YC13-554和YC13-559的遗传相似性系数最大为0.94,确定为近等基因系。而后对这两个材料成熟期的农艺性状进行调查,结果进一步确定了YC13-554和YC13-559为高(中低)含油量近等基因系。2.以含油量差异的近等基因系材料YC13-554和YC13-559为基础材料,利用实时定量PCR的方法,检测转录因子BnWRKY52、BnWRKY16在种子成熟过程中的基因表达量。结果显示,在种子成熟的不同时期BnWRKY52在油菜YC13-554和YC13-559中的表达均呈现上升趋势,且在油菜YC13-559中表达量比在YC13-554中高;WRKY16在油菜YC13-554和YC13-559的表达均只出现一个较大的表达峰,在油菜YC13-559中的表达峰出现早于油菜YC13-554中。据以上结果表明,转录因子WKY52和WRKY16与油菜油分合成的调控有关。为验证所得结果,选择了三个已确定的与油菜含油量有关的关键酶的基因二酰基甘油酰基转移酶(DTAG)、乙酰辅酶A羧化酶、Wril,检测其表达量,并使BnWRKY52、BnWRKY16表达量与之进行对比。结果显示,两者之间表达模式相似。3.以近等基因系材料YC13-554和YC13-559为基础材料,利用实时定量PCR技术检测转录因子BnWRKY52、BnWRKY16在油菜种子萌发过程中的基因表达量。结果显示,在种子萌发的不同时期,BnWRKY52、BnWRKY16在油菜YC13-554与油菜YC13-559中的表达均有两个表达峰,分别在幼苗突破种皮与长出第一片真叶的时期,且在YC13-559中比YC13-554中起始表达早。推测BnWRKY52与BnWRKY16参与油菜种子萌发过程中油脂的代谢调控。
[Abstract]:Rape is the main oil crop planted in China, which plays an important role in the development of our national economy.Improving the oil content of rapeseed has been the primary task of rapeseed breeding. WRKY transcription factor plays an important role in the plant regulatory network as a regulatory factor, and participates in a series of regulation processes such as plant growth and development, stress resistance, disease resistance and so on.Two WRKY transcription factors, BnWRKY52 and BnWRKY52, which are related to oil content, were obtained by sequencing and analysis of transcriptome.The details are as follows: 1.Using NJ9 high oil strain as the basic material, a pseudo isogenic line with different oil content was constructed by continuous self-crossing.Nine pseudo isogenic lines were amplified by 156 pairs of SSR primers from the common database of Brassica Brassica DB. 32 pairs of primers with clear bands and good reproducibility were selected for near-isomorphism analysis.Based on the results of 32 pairs of SSR primer PCR amplification, genetic and similarity coefficients were estimated and cluster analysis was used to identify the near isogenic lines. The results showed that the genetic similarity coefficients of the 9 candidate lines ranged from 0.85 to 0.94.The genetic similarity coefficient between YC13-554 and YC13-559 was 0.94, which was identified as a near isogenic line.Then the agronomic characters of the two materials at maturity were investigated. The results showed that YC13-554 and YC13-559 were high (medium and low) oil content near isogenic lines.The gene expression of transcription factor BnWRKY52 and BnWRKY16 during seed maturation was detected by real-time quantitative PCR based on YC13-554 and YC13-559.缁撴灉鏄剧ず,鍦ㄧ�嶅瓙鎴愮啛鐨勪笉鍚屾椂鏈烞nWRKY52鍦ㄦ补鑿淵C13-554鍜孻C13-559涓�鐨勮〃杈惧潎鍛堢幇涓婂崌瓒嬪姡�

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