Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement, Institute of Vegetable Crops, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, People's Republic of China.
College of Resources and Environmental Sciences, China Agricultural University, Beijing, 100083, People's Republic of China.
Planta. 2019 May;249(5):1599-1615. doi: 10.1007/s00425-019-03108-3. Epub 2019 Feb 15.
Alternative splicing (AS) events were identified and verified in cabbage by comparative transcriptome analysis. The corresponding markers were developed and the germplasm resources were identified. Alternative splicing (AS) is a central regulatory mechanism that greatly contributes to plant gene expression and transcriptome diversity. A large body of evidence has shown that AS complexity is relevant for plant development, evolution, complexity, and adaptation. Both insertion/deletion (InDel) and single nucleotide polymorphism (SNP) are typically co-dominant inheritance markers and have abundant polymorphisms. These have been widely used for marker-assisted selection, genetic mapping, and germplasm identification in plants. However, little is known about the molecular mechanisms underlying AS events and the development of markers including SNP and InDel from the cabbage transcriptome. In this study, three cabbage transcriptome datasets were collected and aligned to the cabbage reference genome to analyze AS events and marker development. 31,524 AS events were identified from three cabbage genotypes, accounting for 20.8% of the total cabbage genes. Alternative 3' splice site donor (A3SS) was the most frequent type of the four main AS events in cabbage. 70,475 InDels and 706,269 SNPs were identified with average frequencies of 1 InDel/6.9 kb and 1 SNP/0.7 kb, respectively. 71,942 potential SSRs were identified in 53,129 assembled unigenes with a density of 1 SSR/6.8 kb. The ratio of SNPs with synonymous/non-synonymous mutations was 1:0.65. 142 InDels and 36 SNPs were randomly selected and validated via Sanger sequencing and polymorphism was found among 66.2% of the InDels and 78.6% of the SNPs. Furthermore, 35 informative InDel markers were successfully used for genetic diversity analysis on 36 cabbage accessions. These results facilitate understanding of the molecular regulation mechanism underlying AS events in cabbage. They also provide molecular marker resource data for genetic mapping construction and germplasm identification, and facilitate the genetic improvement of cabbage via breeding.
白菜中通过比较转录组分析鉴定和验证了可变剪接（AS）事件。开发了相应的标记，并鉴定了种质资源。可变剪接（AS）是一种重要的调控机制，极大地促进了植物基因表达和转录组多样性。大量证据表明，AS 复杂性与植物发育、进化、复杂性和适应性有关。插入/缺失（InDel）和单核苷酸多态性（SNP）通常是共显性遗传标记，具有丰富的多态性。这些标记已广泛用于植物的标记辅助选择、遗传图谱构建和种质资源鉴定。然而，对于 AS 事件的分子机制以及包括 SNP 和 InDel 在内的标记从白菜转录组中的发展知之甚少。在这项研究中，收集了三个白菜转录组数据集，并与白菜参考基因组进行比对，以分析 AS 事件和标记的开发。从三个白菜基因型中鉴定出 31524 个 AS 事件，占白菜总基因的 20.8%。白菜中四种主要 AS 事件中，最常见的类型是可变 3' 剪接位点供体（A3SS）。鉴定出 70475 个 InDel 和 706269 个 SNP，平均频率分别为 1 个 InDel/6.9 kb 和 1 个 SNP/0.7 kb。在 53129 个组装的非基因中鉴定出 71942 个潜在 SSR，密度为 1 SSR/6.8 kb。同义/非同义突变 SNP 的比例为 1:0.65。随机选择 142 个 InDel 和 36 个 SNP 进行 Sanger 测序验证，发现 66.2%的 InDel 和 78.6%的 SNP 存在多态性。此外，35 个信息性 InDel 标记成功用于 36 个白菜种质资源的遗传多样性分析。这些结果有助于理解白菜中 AS 事件的分子调控机制。它们还为遗传图谱构建和种质资源鉴定提供了分子标记资源数据，并通过育种促进了白菜的遗传改良。
