日本落叶松生长及材性相关基因关联遗传研究

发布时间：2018-03-02 16:37

  本文选题：日本落叶松　切入点：群体结构　出处：《中国林业科学研究院》2016年博士论文　论文类型：学位论文

【摘要】：木材产量和品质改良是林木育种研究的关键科学问题,木材形成是一个多基因控制的复杂过程,深入解析木材形成的关键因子和调控机理,有助于借助分子设计手段加快育种进程。日本落叶松(Larix kaempferi)是我国东北、华北和西南亚高山地区的重要针叶造林树种之一,也是我国主要针叶纸浆材用树种。本研究以17年生日本落叶松种源/家系试验林为研究对象,选取具有代表性的603个基因型个体建立关联群体,在分析表型多样性、遗传多样性、核苷酸多样性及连锁不平衡水平基础上,开展100个SNP标记和20对SSR标记与21个生长、形质、木材纤维及木材化学等表型性状的关联研究,旨在筛选出与木材生长和品质相关联的功能标记,为日本落叶松分子设计育种和标记辅助育种奠定基础。主要研究结论如下:1、日本落叶松关联群体表型多样性分析显示:日本落叶松具有丰富的表型变异,表型变异系数为3.08%-54.84%,不同性状的表型变异大小为:生长性状纤维性状形质性状木材化学性状。关联群体各性状间及与地理环境因子间的相关性分析共检测到134对显著或极显著正负相关关系。主成分分析有效验证了性状间的相关性,并将21个表型性状降维成10个主成分。关联群体中6个自然群体的表型方差分析显示,除冠幅和早晚材微纤丝角的群体间差异不显著外,其他性状均存在显著或极显著差异;表型变异系数及表型分化系数分析表明日本落叶松自然群体具有丰富的表型变异;表型性状聚类分析将6个自然群体划分为两个类群。关联群体和人工群体表型性状的比较分析显示,大部分表型性状平均值均较自然群体显著提高,但部分木材纤维性状(如晚材纤维长和纤维长宽比等)平均值反而降低。2、生长及材性相关基因的SNP分析表明日本落叶松具有较高的核苷酸多样性水平。在扩增的27301 bp序列中共发现1698个SNP位点,SNP发生频率为1/16;总核苷酸多样性水平θw和πT变化范围分别为0.0035-0.2167和0.00224-0.03045。中性检验结果表明大部分候选基因在进化过程中主要受纯化选择作用。进一步的LD分析显示对日本落叶松开展基于候选基因的关联研究是可行的。20个候选基因中仅有Lac2和AE2基因在1500 bp时,R2值才降至0.1以下,其余18个基因的LD在R20.1,核苷酸序列长度约为250 bp-1000bp时就已消失。定量PCR分析表明20个生长及材性相关基因在日本落叶松根、成熟木质部、未成熟木质部、形成层、韧皮部和叶片等6个组织均有表达,其中生长相关基因在成熟叶片表达量最高,材性相关基因在成熟木质部和未成熟木质部高表达。3、利用SSR和SNP标记对日本落叶松自然群体进行遗传多样性分析,鉴于日本落叶松本身的生物学特性(物候差异、花粉流和种子流)、分布区域的复杂地形变化及生境片段化影响,日本落叶松在针叶树种中具有中等水平的遗传多样性(He=0.626);人工群体遗传多样性分析显示经过高强度的生长性状人工选择后,子代依然保存着较高的遗传多样性(He=0.613)。分子方差和遗传分化系数分析表明日本落叶松遗传变异主要以群体内变异为主(遗传距离:0.05-0.093)。基于SSR、SNP和SSR+SNP的自然群体遗传结构分析可将6个自然群体划分为2个类群。以自然群体结构划分结果为基础,采用SSR+SNP的标记组合可将关联群体划分为2个类群。类群Ⅰ主要由草津和浅间等自然群体构成,类群Ⅱ主要由富士和日光等自然群体及部分人工群体构成。4、以关联群体的遗传结构矩阵(Q矩阵)和亲缘关系矩阵(K矩阵)作为协变量,采用MLM模型进行关联研究。20个SSR位点中只有BC241与3年生胸径和材积显著关联。在20个生长及材性相关基因的100个候选SNP中,共检测到25个SNP位点与19个表型性状显著关联,对表型性状的解释率为1.09%-4.75%。生长相关基因中共有10个SNP位点与13个表型性状显著关联,其中AE基因家族中的AE2和AE3基因与部分形质性状和木材纤维性状显著关联。10个木材性状相关基因中,共检测到12个SNP位点与12个表型性状显著关联,对表型性状的解释率为1.09%-4.75%。转录及调控因子中,mi R397a有1个位点与早材微纤丝角和4个晚材性状(晚材壁腔比、晚材纤维长、晚材纤维宽和晚材率)显著关联;MYB转录因子中有两个位点(MYB1-337和MYB1-392)与密度显著关联。20个候选基因中,AE2、AE3和POD等3个基因与生长、形质、早晚材纤维性状和木材化学性状均显著关联,可作为木材性状改良的重点关注对象;MIR397a、CESA2和CSL基因主要与晚材发育显著相关,可作为晚材性状改良的重要标记;MYB转录因子主要与密度显著关联,是进行密度改良的重要标记。
[Abstract]:Yield and quality improvement of wood is the key scientific issues of forest breeding, wood formation is a complex process control more than one gene, the key factor and the regulation mechanism of wood formation in-depth analysis, help to speed up the breeding process by means of molecular design. Japanese Larch (Larix kaempferi) is in the northeast of China, one of the most important coniferous species north and South West Mountain area, is also the main coniferous pulpwood in China with tree species. In this study 17 years of Japanese Larch provenance / family plantation as the research object, selects 603 representative base as individuals to establish related groups, in the analysis of phenotypic diversity, genetic diversity, nucleotide diversity and the level of linkage disequilibrium based on the development of 100 SNP markers and 20 SSR markers and 21 morphological, growth, association of wood fiber and wood chemical phenotypic traits, designed to filter out With the growth and quality of the wood functional marker, lay the foundation for the design of Japanese Larch molecular breeding and marker assisted breeding. The main conclusions are as follows: 1. The diversity of Japanese Larch populations association analysis showed that Japanese Larch had rich phenotypic variation, phenotypic variation coefficient is 3.08%-54.84%, the size of phenotypic variation in different traits: growth characters of fiber quality traits traits of wood chemical traits. The correlation analysis between the characters and related groups and geographic environmental factors were detected on 134 significant or extremely significant positive and negative correlation. Principal component analysis verified the correlation among traits, and 21 phenotypic traits of dimensionality reduction into 10 principal components of phenotypic variance 6. A group of natural population association analysis showed that except the crown width and earlywood microfibril angle difference between groups was not significant, other traits were significant or extremely Significant difference; phenotypic variation coefficient and phenotypic differentiation coefficient analysis showed that natural populations of Japanese Larch had rich phenotypic variation of phenotypic traits; cluster analysis of 6 natural populations were divided into two groups. Comparison analysis showed that the correlation group and artificial population phenotypic traits, some phenotypic traits were the average natural population increased significantly, but some wood fiber characteristics (such as wood fiber length and fiber length to width ratio) the average value of.2 decreased, the growth and material related gene SNP analysis showed that the nucleotide of Japanese Larch has higher diversity level. 1698 SNP loci in 27301 BP amplification of the SNP frequency is 1/16, the total nucleotide diversity; the level of W and theta PI T range were 0.0035-0.2167 and 0.00224-0.03045. neutral test results showed that most of these genes is mainly affected by purifying selection in evolution The role of LD. Further analysis showed that the development of Japanese Larch association studies of candidate genes based on Lac2 and AE2.20 are the only viable candidate genes in BP gene in 1500 when the R2 value was below 0.1, the remaining 18 of the LD gene in R20.1, nucleotide sequence length is about 250 bp-1000bp when it has quantitative disappear. PCR analysis showed that 20 genes related to growth and wood properties in Japanese Larch roots, mature xylem, immature xylem, cambium, phloem and leaf expression in 6 tissues, the expression of growth related genes in mature leaves was the highest material related genes in mature xylem and immature xylem of high expression of.3 and to analyze the genetic diversity of natural populations of Japanese Larch using SSR and SNP markers, in view of the biological characteristics of Japanese Larch itself (phenology, pollen flow and seed flow), distribution area of complex terrain And the effects of habitat fragmentation, genetic diversity of Japanese larch is in the middle level of the coniferous species (He=0.626); population genetic diversity analysis showed that after artificial artificial selection of high strength growth traits, the offspring still have high genetic diversity (He= 0.613). The molecular variance and the coefficient of genetic differentiation analysis showed that Japanese Larch genetic variation of the main body of group variation (genetic distance: 0.05-0.093). Based on the analysis of SSR, SNP and SSR+SNP in the natural population genetic structure of 6 natural populations can be divided into 2 groups. The natural group structure based on the results of using SSR+SNP markers can be divided into 2 groups of related groups. Group I consists of cyanazine and the shallow natural population composed of group II mainly by Fuji and sun and other natural populations and groups constitute part of the artificial.4, with genetic structure related groups Matrix (Q matrix) and relationship matrix (K matrix) as a covariate, using MLM model to study the correlation between.20 BC241 and SSR loci in only 3 years. Significant correlation between DBH and volume in the 100 candidate SNP 20 growth and material related genes, 25 SNP and 19 significant correlation between phenotypic traits were detected on phenotypic traits accounted for 1.09%-4.75%. of growth related genes in a total of 10 SNP loci and 13 phenotypic traits were significantly associated, which the AE family of genes AE2 and AE3 genes and some quality traits and wood fiber traits significantly associated.10 wood traits related genes, 12 SNP loci with 12 phenotypic traits significant correlation were detected on phenotypic traits accounted for 1.09%-4.75%. transcription and regulation factor, MI R397a 1 loci and earlywood microfibril angle and 4 latewood traits (latewood ratio of wall to cavity, late wood fiber length, latewood Fiber width and latewood percentage) was significantly associated with two sites; MYB transcription factors (MYB1-337 and MYB1-392) and density significantly associated candidate genes.20, AE2, AE3 and POD form 3 gene and growth traits and wood chemistry, early, late wood fiber traits were significantly correlated, as the focus of wood properties improvement the object of attention; MIR397a, CESA2 and CSL genes were significantly related to late wood development, can be used as an important marker of latewood trait improvement; MYB transcription factors significantly associated with density, is an important marker of density improvement.

【学位授予单位】：中国林业科学研究院
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S791.223
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本文编号：1557315