黄瓜果实长度性状QTL定位及候选基因筛选

发布时间：2018-01-08 07:10

本文关键词：黄瓜果实长度性状QTL定位及候选基因筛选　出处：《南京农业大学》2016年博士论文　论文类型：学位论文

【摘要】：黄瓜(Cucumis sativus L.)是具有重要经济价值的蔬菜作物,在世界范围内广泛栽培。果实的大小和形状是黄瓜品质评价、分类以及市场定级的主要参考指标,同时也是商业品质育种的重要依据之一。黄瓜果实的大小和形状受外在环境因素与内在遗传机制的影响,与果实长度、果实横径或果形指数有关;研究表明黄瓜果实大小主要受到果实长度的影响。然而,控制黄瓜果实长度的分子机制尚不明确。近年来,随着新一代测序技术与生物信息学的不断发展,分子生物学与基因组学的研究方法也随之改变;在数量性状(QTL)定位方面,开发大量的SNP标记并应用于遗传图谱的构建成为可能,而且在不构建遗传图谱的情况下,也能快速识别控制作物数量性状的主效QTLs。本研究利用果实大小和形状差异较大的两个自交系亲本构建分离群体,结合最新的SLAF-seq和QTL-seq技术,对黄瓜子房长度、商品瓜长度和成熟瓜长度进行了系统的QTL定位研究,并筛选到与果实长度相关的候选基因,主要结果如下:1.黄瓜果实发育的生长指数统计与切片观察利用自交系材料NC76,通过对开花前子房和授粉受精后黄瓜果实的果实长度和横径性状数据的统计以及石蜡切片观察,可将黄瓜果实发育大致分为三个阶段;普通黄瓜一般具有三心室,三心室黄瓜的果形指数主要受果实长度影响,受果实横径影响较小;对子房长度、商品瓜长度和成熟瓜长度性状数据的分析发现,三者具有较高的相关性,因此通过黄瓜子房长度可以预测商品瓜和成熟瓜的果实长度。2.黄瓜高密度遗传图谱构建与商品瓜和成熟瓜果实长度QTL定位利用果实长度差异较大的自交系材料CC3和NC76为亲本,杂交自交获得148株F2群体,对亲本及F2单株进行简化基因组测序(SLAF测序),开发1800个SNP标记,并构建了黄瓜首张高密度SNP遗传图谱,该图谱总长度为890.79cM,标记的平均间距为0.5 cM。利用1800个SNP标记锚定了黄瓜基因组183个scaffolds,共计168.9 Mb,结合荧光原位杂交技术(FISH)发现了基因组scaffold000063_5和scaffold000063_1顺序拼接错误。利用F3家系群体,对商品瓜和成熟瓜果实长度进行性状统计(2013秋和2014春),并利用MapQTL和Rqtl两种软件对商品瓜长度和成熟瓜长度进行QTL定位分析。共计发现8个与商品瓜和成熟瓜长度相关的一致性QTLs,分布在1、3、4、6号染色体上。3.利用QTL-seq技术对子房长度的QTL分析通过单粒传方式获得F7重组自交系群体,利用RIL群体开花当天的子房长度性状数据,挑选极端长度的单株并进行混池测序,结合QTL-seq技术,分别利用两个亲本重测序数据作为参考序列,计算SNP-index,发现了 6个与子房长度相关的QTLs。4.黄瓜果实长度候选基因筛选利用番茄和拟南芥中已经发现的与果实大小相关的基因序列信息,通过BLAST比对分析,发现了黄瓜基因组中87个果实大小同源基因;根据对子房长度、商品瓜长度和成熟瓜长度的QTL定位分析结果,总结得出与黄瓜果实长度相关的主效一致性QTL位点FL3.2;有6个果实大小同源基因分布在FL3.2内,定量分析和基因功能注释推测基因Csa3M812740.1是控制果实长度的候选基因。
[Abstract]:Cucumber (Cucumis sativus L.) is the important economic value of vegetable crops, widely cultivated in the world. The size and shape of cucumber fruit quality evaluation, index classification and grading of the market at the same time, and is one of the important bases of commercial quality breeding of cucumber fruit. The size and shape of the affected by the external environment and internal factors the genetic mechanism, and the fruit length, fruit diameter and fruit shape index of cucumber fruit size; research shows that mainly affected by the length of fruit. However, the molecular mechanisms that control the length of cucumber fruit is not clear. In recent years, with the continuous development of a new generation of sequencing technology and bioinformatics, molecular biology and research methods genomic changes in the number of characters; (QTL) positioning, construction can be developed and applied to a large number of SNP markers in genetic map, but not in structure The construction of genetic map case, also can quickly identify the main effect QTLs. control of crop traits based on fruit size and shape difference of two inbred lines to construct larger separation groups, combined with the latest SLAF-seq and QTL-seq technology, the cucumber ovary length, fruit length and fruit length of mature QTL positioning system, and screened for candidate genes associated with fruit length, the main results are as follows: the growth index statistics and slice observation using NC76 1. inbred lines of cucumber fruit development, based on the pre anthesis and ovary after pollination of cucumber fruit fruit length and diameter traits data statistics and paraffin section observation, the cucumber the fruit development can be divided into three stages; the ordinary cucumber generally has three ventricle, three ventricle of cucumber fruit shape index is mainly affected by fruit length, fruit diameter is less affected by the; The ovary length, fruit length and fruit maturity analysis length trait data found that three have high correlation, so the length of cucumber ovary can predict the fruit length of cucumber.2. to construct high density genetic map of melon and watermelon mature commodity and commodity melon and mature melon fruit length length of QTL positioning using different inbred lines materials CC3 and NC76 as parental strains, 148 strains of F2 hybrids were selfed to obtain groups of parents and F2 simplified plant genome sequencing (SLAF sequencing), developed 1800 SNP markers, and constructed the first cucumber SNP high density genetic map, the total length of the map was 890.79cM, the average distance between markers was 0.5 cM. by 1800 a SNP anchor marker of cucumber genome of 183 scaffolds, a total of 168.9 Mb, combined with fluorescence in situ hybridization (FISH) genomic scaffold000063_5 and scaffold000063_1 sequence splicing errors were found. F3 family groups, the character statistics of commodity melon and ripe melon fruit length (2013 autumn and spring 2014), and QTL analysis of fruit length and ripe melon length by MapQTL and Rqtl two software. A total of 8 commodities and found the melon and ripe melon length is related to the consistency of QTLs distribution in the 1,3,4,6 on chromosome.3. by using QTL-seq technology on the length of the QTL F7 analysis of recombinant inbred lines by single seed descent, flowering data length. The traits in the RIL population, extreme length and plant selection of mixed pool sequencing, combined with QTL-seq technology, using two parent resequencing data as a reference sequence, calculate SNP-index, found 6 related to the length of the ovary QTLs.4. candidate gene screening using the length of cucumber fruit and fruit size related gene sequence information has been found in tomato and arabidopsis, Through BLAST analysis, found the cucumber genome 87 homologous genes according to fruit size; ovary length, length and location of QTL fruit ripe melon length analysis results, summarize the related main effect of cucumber fruit length consistency QTL locus FL3.2; 6 fruit size distribution of homologous genes within the FL3.2. Quantitative analysis and gene annotation that Csa3M812740.1 gene is a candidate gene for controlling the length of fruit.

【学位授予单位】：南京农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S642.2

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