基于MSAP技术的刺参选育群体基因组表观与序列遗传多样性分析

Evaluation of epigenetic and genome sequence diversity in sea cucumber Apostichopus japonicus selected population based on MSAP technology

关键词:  刺参  MSAP  DNA甲基化  选育群体  遗传多样性

Key words:  Apostichopus japonicus  MSAP  DNA methylation  Breeding population  Genetic diversity

摘要:

抗逆选育引起的遗传变化不仅源于DNA序列的变化，也有来自于表观层面的修饰改变。为探究刺参(Apostichopus japonicus)耐高温新品系育种过程中的选育基础群体与选育群体的遗传多样性，运用MSAP技术分析了选育基础群体F、选育F1代和选育F4代的基因组遗传多样性。结果显示，10对引物获得的806个位点中，多态性位点为698个，多态性百分比达到86.60%；基于非甲基化位点的遗传分析，选育F4代香农多态性指数为0.3981，Nei基因多样度为0.2264；基于甲基化敏感位点分析，选育F4代香农多态性指数为0.5873，Nei基因多样度为0.2598，均高于基础群体；表观遗传多样性均大于非甲基化位点变异产生的序列遗传多样性，表明表观变异出现频率高于序列遗传变异。MSAP甲基化模式分析显示，选育F1和F4代经过选育后获得了一些甲基化水平和模式的改变，说明经温度胁迫选育，改变了刺参群体的基因组的甲基化状态。选育F4代获得的类型Ⅱ的条带数最多，为161条，明显高于未选育刺参，为选育获得表观遗传特征。研究结果从遗传物质基础角度揭示了选育群体的遗传改变与进展，可为抗逆新品种选育中的表观遗传研究提供参考。

Abstract:

Apostichopus japonicus is a species of economic importance cultured in northern China. In the past 10 years, the scale of the A. japonicus culture has expanded due to increased market demand for products. However, several challenges, such as germplasm degradation and the lack of stress-resistant varieties, have emerged in A. japonicus culture. Breeding new varieties is one way to overcome these challenges. New strains (F1 and F4) of A. japonicus with high temperature resistance were obtained by domestication and screening under environmental stress conditions. The genetic changes caused by breeding under environmental stress conditions not only originated from changes in the DNA sequence, but also from changes due to epigenetic modifications by bisulfite sequencing. To explore the genetic diversity of the selected populations (F1 and F4) and the control population (F) which was not subjected to temperature stress of A. japonicus, the genetic diversity of control population, selected population F1, and selected population F4 were analyzed using methylation-sensitive amplification polymorphisms (MSAP). The results showed that 698 loci were polymorphic among the 806 loci obtained by 10 pairs of primers, and the percentage of polymorphism was 86.60%. Based on the genetic analysis of non-methylated loci, the Shannon polymorphism index of the F4 was 0.3981 and Nei gene diversity was 0.2264. Based on the analysis of methylation sensitive sites, the Shannon polymorphism index of the F4 was 0.5873, and Nei gene diversity was 0.2598, both of which were higher than in the parent population. Moreover, the epigenetic diversity was higher than the sequence genetic diversity caused by variation in non-methylated loci, indicating that the frequency of epigenetic variation was higher than that of sequence genetic variation. Analysis of the MSAP methylation patterns revealed some changes in the methylation levels and patterns in the selected F1 and F4 after breeding, which indicated that the genome methylation status of the A. japonicus population was changed by breeding under temperature stress. The number of type Ⅱ bands in the F4 was 161, which was significantly higher than that of control population of A. japonicus and may have acquired epigenetic characteristics during breeding. Collectively, our results revealed the genetic changes and progress of the breeding population from the perspective of a genetic material basis and provide a reference for the study of epigenetics in the breeding of new stress resistant varieties.