应用简化基因组技术对富民枳遗传多样性检测

Genetic Diversity Analysis of Poncirus polyandra by Reduced-representation Sequencing Technique

富民枳是云南特有的极小种群野生植物,野生资源仅剩15株。通过二代测序技术,对其进行简化基因组测序,开发一批特异性高的单核苷酸多态性标记,分析天然种群和回归种群的遗传多样性。经过遗传变异检测,共获得SNP位点491 189个,通过样品最低测序深度>2,样品缺失率<0.5、次要基因型频率(MAF)>0.05筛选以后,得到有效SNP位点41 077个。基于过滤后的SNP,运用生物信息学分析方法,对富民枳完成了群体的遗传多样性分析,研究分析了富民枳天然种群和回归种群的私人等位基因数目、平均观测杂合度(Ho)、平均期望杂合度(He)、核苷酸多样性(π)和平均近交系数(FIS)5个遗传多样性参数。结果表明,富民枳现存植株的遗传多样性较高,具有很高的遗传资源保存价值,但是回归种群的Ho值、He值、π值分别为0.398 3、0.291和0.330 6,分别仅是天然种群遗传多样性的87%、78%和73%,并未达到90%以上遗传多样性的标准。

Poncirus polyandra is a plant species with extremely small populations endemic to Yunnan Province with only 15 wild plants left. The recently developed technology restriction-site associated DNA sequencing(RAD-seq), which is from next-generation sequencing(NGS), was used to develop a batch of highly specific single nucleotide polymorphism(SNP) markers and analyze genetic structure and genetic diversity of existing populations. After detection of genetic variation, a total of 98 498 SNPs were got. Under these conditions, the minimum sequencing depth of samples >2, sample missing rate <0.5 and MAF>0.05 screening from all 491 189 SNPs, 41 077 effective SNPs were obtained. Genetic diversity analysis of P. polyandra was analyzed using bioinformatics method with screening SNPs. The genetic diversity parameters of P. polyandra natural population and reintroduction population, involving private allele number, the nucleotide diversity(π), observed heterozygosity(Ho), expected heterozygosity(He) and inbreeding coefficient(FIS), were elucidated. P. polyandra had higher genetic diversity. Therefore, preserving genetic resources was of very high value. However, Ho value, He value and π value of the reintroduction population were 0.398 3, 0.291 0 and 0.330 6, respectively, which were only 87%, 78% and 73% of the genetic diversity of natural population, and did not meet the standard of genetic diversity, which was more than 90%.