桃树“亲—子”代突变率与重组率的研究

发布时间：2018-01-21 20:15

本文关键词： 突变同源重组桃树杂合度世代周期　出处：《南京大学》2017年博士论文　论文类型：学位论文

【摘要】：突变和同源重组是生物体基因组遗传多样性的主要来源,也是生物演化过程中不可或缺的重要步骤。目前,科学家们已经对多种生物(酵母,果蝇,小鼠,人,拟南芥、水稻等)的突变或同源重组都进行了相关报道研究,但系统性的研究主要集中在个别模式物种,对于多年生木本植物,则仅局限于基因组的某些特定区段,或者依赖于有限的分子标记位点,很容易受到遗传背景差异等各种因素的影响,缺少系统、精确和可靠的评估。另外,很多研究表明,不同物种或者同一物种内的不同个体,其突变率与重组率与其世代周期、参与DNA修复系统的效率和精度、杂交方式以及基因组的杂合度均存在一定的相关性。而这些研究结果在不同物种当中的普适性还亟需更多的数据来支持和证明。本研究采用二代测序的方法,对多年生木本桃树的亲本-后代群体的一次减数分裂的突变率和重组率进行了全基因组范围内的精确评估,并与一年生模式植物拟南芥进行比较,探究影响植物体突变和重组的主要因素。本研究构建了3组桃树种内和种间组合,并对每一组F1杂交个体及其F2自交后代个体进行二代测序。通过生物信息学分析,从3组共63个F2自交后代中检测到240个点突变和46个短片段的插入缺失突变。结果表明,杂合度最高的种间杂交组合Ⅲ的突变率大约是杂合度较低的种内杂交组合Ⅰ突变率的1.8倍(BM检验,p=2.22×10-5碱基突变和p=0.0064插入缺失突变)。而不同组之间突变的特征,点突变与插入缺失突变的比例却并无明显差异。其次,在一个世代周期内,桃树种内杂交组合一次减数分裂的突变率与一年生植物拟南芥相差无几。这与桃树的世代时长约为拟南芥世代时长的10～20倍形成鲜明对比,表明单位时间内的桃树的突变率比拟南芥慢了约1个数量级,从而直接验证了世代周期对于物种进化速率的贡献。桃的重组率评估中,种内组合Ⅰ的24个F2样品中共检测到286个交叉互换,相当于每次减数分裂每个样本平均11.92次交叉互换或2.64cM/Mbp,与之前研究结果一致。作为比较,我们利用四分孢子分析,精确评估拟南芥中一次减数分裂约发生11.11次交叉互换,重组率约为4.66cM/Mbp。可以看出,桃中每次减数分裂每个样本的交叉互换率低于一年生双子叶植物拟南芥(4.0cM/Mbp),同时也低于一年生单子叶植物水稻(4.53cM/Mbp)。该结果验证了桃树与其他多年生木本植物(如苹果、梨、葡萄、橡木和胡桃木,0.63～2.5cM/Mbp)相对于一年生植物(如拟南芥和水稻)的低重组率。除此之外,桃树的重组率缺乏与突变明显的相关性,可能与栽培桃受到的驯化及强烈的人工选择有关。最后,与一年生植物类似,桃树存在明显的重组冷点和重组热点,且重组热点发生区域的规律与非PRDM9物种一致。本研究的创新之处在于,第一次用全基因组测序的方式直接评估了多年生木本桃树一次减数分裂的突变率与重组率,填补了全基因组范围内多年生木本植物突变率与重组率精确评估的空白。通过与拟南芥的突变率与重组率进行比较,系统性检验了突变、重组与世代周期长短的关系,并进一步验证了杂交方式或者杂合度对突变率的影响,为物种演化研究提供了新的材料,也为突变与重组机制的深入发掘奠定了基础。
[Abstract]:Mutation and homologous recombination is a major source of genetic diversity of organisms, is an important step in the process of biological evolution. At present, scientists have to many organisms (yeast, Drosophila, mouse, human, Arabidopsis, rice) mutation or homologous recombination have been reported, but the systematic research mainly in the mode of individual species, the perennial woody plant, certain section is limited to the genome, or rely on molecular markers in limited, easily affected by differences in the genetic background of various factors of the lack of systematic, accurate and reliable evaluation. In addition, many studies have shown that different individuals of different species or within the same species, the mutation rate and recombination rate and generation time, efficiency and accuracy in DNA repair system, hybridization and gene heterozygosity there were certain groups Correlation. These results in different species of universality also need more data to support and verify. This research adopts the two generation sequencing, the perennial woody peach parent offspring group of a meiotic division rate of mutation and recombination rates were all based on accurate assessment within the group the annual, and compared with the model plant Arabidopsis thaliana, explore the main factors influencing plant somatic mutation and recombination. This study established 3 groups of peach tree intraspecific and interspecific combinations, and the two generation sequencing of each group of F1 hybrids and F2 inbred offspring. Through bioinformatics analysis, detection from 3 groups a total of 63 F2 progenies to 240 point mutations and 46 short fragments of insertion and deletion. The results showed that the highest heterozygosity of interspecific hybrids of the mutation rate is about intraspecific crosses of sudden low heterozygosity 1.8 times the rate of change (BM test, p=2.22 * 10-5 mutation and p=0.0064 mutation and insertion mutation). Between different groups, point mutation and insertion mutation ratio has no obvious difference. Secondly, in a generation period, peach intraspecific crosses a meiotic mutation rate and the annual plant Arabidopsis thaliana. The peach and not much difference between generation time is approximately the length of the 10 generations of Arabidopsis thaliana to 20 times in stark contrast to that mutation in unit time and rate of peach thaliana slow about 1 orders of magnitude, which directly verify the contribution period for the evolution of species generation rate. The assessment rate of peach the reorganization, I combined within 24 F2 samples were detected in 286 crossover, each equivalent to an average of 11.92 times for each sample of meiotic crossover or 2.64cM/Mbp, consistent with previous reports. As a comparison, we Analysis of four spores using a precise evaluation of Arabidopsis meiosis about 11.11 crossover, recombination rate of about 4.66cM/Mbp. can be seen in each peach meiosis of each sample crossover rate is lower than the annual dicotyledonous plant Arabidopsis thaliana (4.0cM/Mbp), and also lower than the annual monocotyledon rice (4.53cM/Mbp) the results. Verify the peach trees and other woody perennial plants (such as apples, pears, grapes, oak and walnut, 0.63 ~ 2.5cM/Mbp) relative to the annual plant (such as Arabidopsis and rice) the low recombination rate. In addition, the rate of recombination and mutation of peach lacks obvious relevance, domestication and artificial selection may strongly concerned with peach. Finally, similar to the annual peach exist obvious and cold recombination recombination hotspots, and recombination hotspots law consistent with regional non PRDM9 this species. Study on the innovation, for the first time, whole genome sequencing methods to directly evaluate the peach a perennial woody meiotic recombination rate and mutation rate, fill the perennial woody plant genome wide mutation rate blank with recombination rate. Through the comparison of the precise assessment of the rate of mutation and Arabidopsis with recombination rate, system to test the mutation, recombination and generation cycle length, and further verify the hybrid or heterozygosity on influence of mutation rate, provide new material for the evolution of species, but also lay a foundation for the further development and mechanism of recombinant mutant.

【学位授予单位】：南京大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S662.1

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