谷子穗发育调控基因图位克隆与功能分析

发布时间：2018-03-03 17:49

本文选题：谷子　切入点：稀码　出处：《甘肃农业大学》2017年博士论文　论文类型：学位论文

【摘要】：谷子(Setaria italica(L.)P.Beauv.)具有基因组小、重复序列少、生育期短、繁殖系数高等特点,并已完成全基因组测序,正发展成为禾本科新的模式植物。谷子属于稀播作物,穗部性状是决定产量的主要因素。目前,关于调控谷子穗发育基因的研究鲜有报道。本研究以EMS诱变谷子全基因组测序品种Yugu1所获得的穗发育异常突变体siaux1和lp1为对象,图位克隆目的基因并分析其功能,主要获得如下结果:1.与Yugu1相比,siaux1的种子发芽势和发芽率降低,植株茎节数减少、主茎变粗变矮、叶片变长变宽,主穗变长、穗码数减少、小花数减少、结实率降低;最明显的特征是穗码变稀、单穗产量极显著降低。siaux1的突变性状由位于5号染色体编码IAA输入载体(AUX1)的基因Si AUX1(Seita.5G387200)控制;siaux1的Si AUX1基因第四个外显子区发生G→A的突变,导致三联体密码子TGG(UGG)替换为终止密码子TGA(UGA),造成翻译蛋白时在214个氨基酸处提前终止,缺失了C-端的277个氨基酸残基。2.敲除水稻Os AUX1(LOC_Os01g63770)基因突变株的穗密度、侧根密度显著减小;异源超表达Si AUX1基因水稻突变株的穗密度、侧根密度显著增大。50个代表性谷子品种中,7个品种的Si AUX1基因3’UTR区存在38bp的大片段缺失,这些品种的穗长、第一级分枝数、小花数、主穗粒重显著小于其它品种,而穗码密度显著大于其它品种。3.Si AUX1基因能够响应2,4-D的诱导,表达量上调的幅度与2,4-D的处理浓度成正比;Si AUX1基因在生长旺盛的胚芽、心叶、幼穗等幼嫩组织和主根的髓、颖壳的脊等维管组织中高表达,与生长素的分布与运输模式吻合。Si AUX1基因突变会显著下调IAA信号转导途径多个基因在穗部的表达量;siaux1灌浆期生长旺盛的穗部IAA的含量(46.17 mg/g)极显著高于野生型(27.61 mg/g)。谷子AUX/LAX家族有5个基因,只有Si AUX1在穗中高表达,具有调控穗发育的功能;其它4个基因Si AUX2(Seita.3G223500)、Si AUX3(Seita.9G471700)、Si AUX4(Seita.9G288200)、Si AUX5(Seita.8G047400)在不同发育时期的根中高表达。4.与Yugu1相比,lp1的植株主茎变矮,主穗变长、第一级分枝数和第二级分枝数均减少、小花数减少、籽粒数减少;最明显的特征是穗码变稀、单穗产量显著降低,但籽粒变长变宽,千粒重极显著增大。lp1的突变性状由位于2号染色体属于WRKY转录因子家族Ⅰ亚族的基因LP1(Seita.2G369500)控制;lp1的LP1基因第五个内含子末端的碱基发生G→A的突变,形成3个不同于野生型的蛋白翻译提前终止型突变的可变剪切,造成LP1蛋白C-末端第二个WRKY结构域的C2H2锌指结构遭到破坏。5.LP1基因分别在拔节期的茎节、孕穗期的穗、灌浆期的籽粒中高表达,这与其调控株高、穗型和种子大小的功能一致;亚细胞定位显示LP1基因在细胞核中表达,符合转录因子基因的表达特征。谷子的穗发育是受多个基因控制的复杂过程。本研究发现生长素输入载体编码基因Si AUX1和WRKY家族转录因子LP1突变均会导致谷子穗发育异常,表现出明显的稀码特征,且单穗产量显著下降。
[Abstract]:Millet (Setaria Italica (L.) P.Beauv.) has a small genome, repeat, short growth period, high propagation coefficient, and whole genome sequencing has been completed, is developing into a new plant. The model of gramineous crops of millet is rare, panicle traits are the main factors determining yield. At present, there are few reports on on gene regulation millet ear development. In this study, whole genome sequencing EMS mutagenesis millet varieties Yugu1 acquired ear development mutant siaux1 and Lp1 as the object, map based gene cloning and function analysis, the main results are as follows: 1. compared with Yugu1 siaux1, the seed germination potential and germination rate decreased, the stem section the number of plants decreased, stem thick and short blade of variable length and variable width, variable length of main spike, spike number decreased, the number of florets decreased, the seed setting rate decreased; the most obvious feature is the spike code thinning, yield per spike decreased significantly in.Siaux1 process Shaped by degeneration of chromosome 5 IAA encoding the input vector (AUX1) gene Si AUX1 (Seita.5G387200) siaux1 Si control; AUX1 gene mutation of fourth exons of G to A promoter, resulting in three CIS codon TGG (UGG) to replace the termination codon TGA (UGA), caused by protein translation in 214 amino acids at the early termination, the deletion of 277 amino acid residues of.2. C- end of the knockout AUX1 rice Os (LOC_Os01g63770) mutant panicle density, lateral root density decreased significantly; heterologous over expression of Si gene AUX1 in rice mutant panicle density, root density increased significantly.50 representative millet varieties there, a large fragment deletion of 38bp Si AUX1 gene of 7 varieties of 3 UTR region, these varieties of ear length, the first branch number, floret number, main spike grain weight was significantly less than other varieties, and spike code density was significantly greater than the other varieties of.3.Si AUX1 gene in response to 2,4-D By the concentration of 2,4-D was increased and the expression magnitude is proportional to the leaf; Si AUX1 gene in the germ, vigorous growth, such as young spike tissues and root pulp, high expression of tissue glume ridge dimension, consistent with the auxin distribution and transport patterns of.Si mutations in the AUX1 gene can significantly reduce IAA the signal transduction pathways of multiple gene expression in panicle weight; content of strong IAA growth siaux1 ear filling stage (46.17 mg/g) was significantly higher than that of the wild type (27.61 mg/g). Millet AUX/LAX family has 5 genes, only Si high expression of AUX1 in Guangzhou, with regulation of ear development of other functions; 4 genes Si AUX2 (Seita.3G223500), Si AUX3 (Seita.9G471700), Si AUX4 (Seita.9G288200), Si AUX5 (Seita.8G047400) at different developmental stages in the root of the high expression of.4. compared with Yugu1, Lp1 of the plant stem short, main spike becomes longer, the first branch number and level second The number of branches were reduced, the number of florets decreased, grain number decreased; the most obvious feature is the spike code thinning, significant yield per spike decreased, but the grain grow wider, the mutation of LP1 gene traits significantly increased by.Lp1 were located on chromosome 2 belongs to WRKY transcription factor family 1 subfamily (Seita.2G369500) control LP1 gene mutation; fifth introns end Lp1 base G and A, the formation of 3 different from the wild type protein translation termination variable shear mutation, C2H2 zinc caused LP1 protein C- terminal second WRKY domain refers to the destruction of the.5.LP1 gene structure respectively in stem elongation stage. At the booting stage of panicle, high expression in grain filling period, which related to the regulation of plant height, panicle and seed size consistent function; subcellular localization showed that LP1 gene expression in the nucleus, consistent with the expression pattern of transcription factor gene of millet ear hair. Fertility is a complex process controlled by multiple genes. We found that auxin input vector coding gene Si AUX1 and WRKY family transcription factor LP1 mutation will lead to abnormal development of millet panicle, showing obvious sparse code characteristics, and the yield of single spike decreased significantly.

【学位授予单位】：甘肃农业大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S515;Q943.2

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