基于中间锦鸡儿干旱转录组开发SSR标记及遗传多样性研究
发布时间：2018-10-05 14:00
【摘要】：中间锦鸡儿(Caragana intermedia)为豆科、锦鸡儿属多年旱生落叶灌木,生长在海拔900-2400米的固定、半固定沙丘,在我国广泛分布于华北、西北等地,在防风固沙、水土保持方面具有重要作用,而且在饲料、药用和绿化环境等方面也有重要价值。了解中间锦鸡儿的遗传背景,可以为育种材料的选择、研究系统演化及资源合理利用与正确的制定保护策略提供理论依据。分子标记能直接从DNA水平反映群体及个体之间的遗传差异,是研究遗传结构的有力工具。本文基于中间锦鸡儿干旱转录组数据自主开发SSR标记,对内蒙古中部的10个地区进行样品采集并分析其遗传多样性,结果如下:1.基于中间锦鸡儿干旱转录组开发SSR标记:共搜索到404个SSR标记,分布于349条Unigenes序列上,出现频率为14.78%。其中二核甘酸是主要的重复类型,占总SSRs的64.90%;其次是三核苷酸重复,占总SSRs的33.70%;二核苷酸基元以AG/CT和GA/TC为主,占总SSRs的29.45%,三核苷酸基元以AAG/CTT居多,占总SSRs的5.69%。设计并合成了 76对引物,有54对引物能扩增出清晰的条带,并从中筛选出多态性高、重复性好且对柠条锦鸡儿也适用的7对引物。这些引物扩增出等位基因数为3-8个,平均5个;各位点的观察杂合度(Ho)的变化范围是0.108-0.398,平均0.251;期望杂合度(He)范围是0.105-0.495,平均0.268。2.居群的遗传多样性分析:用筛选出的7对多态性引物对10个居群进行遗传多样性分析,结果显示:遗传多样性指标分别为NA=2.547,NE=1.424,HE=0.261,I = 0.445;综合NE、HE、I三类遗传多样性指标对10个居群进行遗传多样性评价,得出的结论是居群总体遗传多样性低,最高的居群为BL(包头市百灵庙镇),最低是居群SZ(乌兰察布市四子王旗)。由NE、HE和I检测居群遗传多样性由高至低依次为BLLCHYHLDGWNGYMAWCSZ。3.居群的遗传分化分析:居群间的遗传距离介于0.0027-0.0722,遗传一致度的范围是0.9304-0.9973,根据遗传一致度对居群进行UPGMA聚类,发现大多数居群以中间锦鸡儿和柠条锦鸡儿为界聚为两类,但有一些居群有交叉现象。居群平均近交系数(Fis)为0.143,说明据居群中有明显的近交现象;基因分化度(Fst)介于0.036-0.164,平均为0.065,表明居群有中等程度的遗传分化;基因流(Nm)很高,平均为3.589,导致居群间的遗传变异较小;对居群的遗传距离和地理距离进行分析,发现二者之间并没有显著的关系(R2=0.023,R2=0.0201);分子方差分析(AMOVA)表明,变异主要发生在居群内个体间。综上分析表明:供试材料的遗传多样性较低,可能是居群采样距离较近或受到人为干预较多,遗传交流大且有明显的近交现象所导致的。
[Abstract]:Caragana Intermediate (Caragana intermedia) is a perennial xerophytic and deciduous shrub of the family Caragana, which grows in fixed and semi-fixed dunes of 900-2400m above sea level. It is widely distributed in North China and Northwest China, and plays an important role in windbreak and sand fixation, soil and water conservation. And in feed, medicinal and green environment and other aspects also have important value. Understanding the genetic background of Caragana intermedia can provide theoretical basis for the selection of breeding materials, the study of system evolution, the rational utilization of resources and the correct formulation of conservation strategies. Molecular markers can directly reflect the genetic differences between populations and individuals from the DNA level and are powerful tools for studying genetic structures. Based on the drought transcriptome data of Caragana intermedia, SSR markers were developed to collect and analyze the genetic diversity of 10 regions in the middle of Inner Mongolia. The results are as follows: 1. Development of SSR markers based on drought transcriptome of Caragana intermedia: 404 SSR markers were identified and distributed on 349 Unigenes sequences with a frequency of 14.78g. Among them, dinucleotide is the main repeat type, accounting for 64.90% of total SSRs, followed by trinucleotide repeat, accounting for 33.70% of total SSRs, dinucleotide motifs are mainly AG/CT and GA/TC, accounting for 29.4545% of total SSRs, and trinucleotide motifs are mostly AAG/CTT, accounting for 5.69% of total SSRs. 76 pairs of primers were designed and synthesized, 54 pairs of primers could amplify clear bands, and 7 pairs of primers with high polymorphism, good reproducibility and suitable for Caragana korshinskii were screened. The number of alleles amplified by these primers ranged from 3 to 8, with an average of 5. The range of observed heterozygosity (Ho) was 0.108-0.398, with an average of 0.251, and the range of expected heterozygosity (He) was 0.105-0.495, with an average of 0.268.2. Population genetic diversity analysis: 10 populations were analyzed with 7 pairs of polymorphic primers. The results showed that the genetic diversity index was NA=2.547,NE=1.424,HE=0.261,I = 0.445, and the genetic diversity of 10 populations was evaluated by NE,HE,I, and the conclusion was that the total genetic diversity of the population was low. The highest population is BL (Baotou Baling Miao Town), and the lowest is SZ (Siziwang Banner of Wulanchabu). The genetic diversity of population detected by NE,HE and I from high to low is BLLCHYHLDGWNGYMAWCSZ.3.. Genetic differentiation analysis of populations: the genetic distance between populations was 0.0027-0.0722, and the range of genetic consistency was 0.9304-0.9973. According to the UPGMA clustering of populations, it was found that most populations were divided into two groups: Caragana intermedia and Caragana korshinskii. But some populations have intersections. The average number of inbred lines (Fis) was 0.143, which indicated that there was obvious inbreeding in the population, the (Fst) of gene differentiation was 0.036-0.164, the average was 0.065, which indicated that the population had moderate genetic differentiation, and the (Nm) of gene flow was very high. The average genetic variation among populations was 3.589, and the genetic distance and geographical distance between populations were analyzed. The results showed that there was no significant relationship between the genetic distance and geographical distance (R2O0.023R20.0201), and the (AMOVA) analysis of molecular variance showed that the variation mainly occurred among the individuals in the population. The results showed that the genetic diversity of the tested materials was low, which might be caused by the close sampling distance of the population or the human intervention, the large genetic exchange and the obvious inbreeding phenomenon.
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;S793.3
本文编号：2253668
[Abstract]:Caragana Intermediate (Caragana intermedia) is a perennial xerophytic and deciduous shrub of the family Caragana, which grows in fixed and semi-fixed dunes of 900-2400m above sea level. It is widely distributed in North China and Northwest China, and plays an important role in windbreak and sand fixation, soil and water conservation. And in feed, medicinal and green environment and other aspects also have important value. Understanding the genetic background of Caragana intermedia can provide theoretical basis for the selection of breeding materials, the study of system evolution, the rational utilization of resources and the correct formulation of conservation strategies. Molecular markers can directly reflect the genetic differences between populations and individuals from the DNA level and are powerful tools for studying genetic structures. Based on the drought transcriptome data of Caragana intermedia, SSR markers were developed to collect and analyze the genetic diversity of 10 regions in the middle of Inner Mongolia. The results are as follows: 1. Development of SSR markers based on drought transcriptome of Caragana intermedia: 404 SSR markers were identified and distributed on 349 Unigenes sequences with a frequency of 14.78g. Among them, dinucleotide is the main repeat type, accounting for 64.90% of total SSRs, followed by trinucleotide repeat, accounting for 33.70% of total SSRs, dinucleotide motifs are mainly AG/CT and GA/TC, accounting for 29.4545% of total SSRs, and trinucleotide motifs are mostly AAG/CTT, accounting for 5.69% of total SSRs. 76 pairs of primers were designed and synthesized, 54 pairs of primers could amplify clear bands, and 7 pairs of primers with high polymorphism, good reproducibility and suitable for Caragana korshinskii were screened. The number of alleles amplified by these primers ranged from 3 to 8, with an average of 5. The range of observed heterozygosity (Ho) was 0.108-0.398, with an average of 0.251, and the range of expected heterozygosity (He) was 0.105-0.495, with an average of 0.268.2. Population genetic diversity analysis: 10 populations were analyzed with 7 pairs of polymorphic primers. The results showed that the genetic diversity index was NA=2.547,NE=1.424,HE=0.261,I = 0.445, and the genetic diversity of 10 populations was evaluated by NE,HE,I, and the conclusion was that the total genetic diversity of the population was low. The highest population is BL (Baotou Baling Miao Town), and the lowest is SZ (Siziwang Banner of Wulanchabu). The genetic diversity of population detected by NE,HE and I from high to low is BLLCHYHLDGWNGYMAWCSZ.3.. Genetic differentiation analysis of populations: the genetic distance between populations was 0.0027-0.0722, and the range of genetic consistency was 0.9304-0.9973. According to the UPGMA clustering of populations, it was found that most populations were divided into two groups: Caragana intermedia and Caragana korshinskii. But some populations have intersections. The average number of inbred lines (Fis) was 0.143, which indicated that there was obvious inbreeding in the population, the (Fst) of gene differentiation was 0.036-0.164, the average was 0.065, which indicated that the population had moderate genetic differentiation, and the (Nm) of gene flow was very high. The average genetic variation among populations was 3.589, and the genetic distance and geographical distance between populations were analyzed. The results showed that there was no significant relationship between the genetic distance and geographical distance (R2O0.023R20.0201), and the (AMOVA) analysis of molecular variance showed that the variation mainly occurred among the individuals in the population. The results showed that the genetic diversity of the tested materials was low, which might be caused by the close sampling distance of the population or the human intervention, the large genetic exchange and the obvious inbreeding phenomenon.
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;S793.3
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