Institute of Horticulture, Henan Academy of Agricultural Sciences, Zhengzhou, China.
College of Life Science, Zhengzhou University, Zhengzhou, China.
PLoS One. 2020 Oct 2;15(10):e0240042. doi: 10.1371/journal.pone.0240042. eCollection 2020.
Kompetitive allele-specific PCR (KASP) is a cost-effective single-step SNP genotyping technology, With an objective to enhance the marker repertoire and develop high efficient KASP-SNP markers in Chinese cabbage, we re-sequenced four Chinese cabbage doubled haploid (DH) lines, Y177-47, Y635-10, Y510-1 and Y510-9, and generated a total of more than 38.5 billion clean base pairs. A total of 827,720 SNP loci were identified with an estimated density of 3,217 SNPs/Mb. Further, a total of 387,354 SNPs with at least 30 bp to the next most adjacent SNPs on either side were selected as resource for KASP markers. From this resource, 258 (96.27%) of 268 SNP loci were successfully transformed into KASP-SNP markers using a Roche LightCycler 480-II instrument. Among these markers, 221 (85.66%) were co-dominant markers, 220 (85.27%) were non-synonymous SNPs, and 257 (99.6%) were newly developed markers. In addition, 53 markers were applied for genotyping of 34 Brassica rapa accessions. Cluster analysis separated these 34 accessions into three clusters based on heading types. The millions of SNP loci, a large set of resource for KASP markers, as well as the newly developed KASP markers in this study may facilitate further genetic and molecular breeding studies in Brassica rapa.
竞争性等位基因特异性 PCR（KASP）是一种经济有效的单步 SNP 基因分型技术。为了增加标记的数量并开发高效的白菜 KASP-SNP 标记，我们对四个白菜加倍单倍体（DH）系 Y177-47、Y635-10、Y510-1 和 Y510-9 进行了重新测序，共产生了超过 3850 亿个清洁碱基对。共鉴定到 827720 个 SNP 位点，估计密度为 3217 SNPs/Mb。此外，选择了总共 387354 个至少与两侧下一个最相邻 SNP 有 30bp 间隔的 SNP 作为 KASP 标记的资源。从这些资源中，使用罗氏 LightCycler 480-II 仪器成功转化为 KASP-SNP 标记的 SNP 位点有 258 个（96.27%）。其中，221 个（85.66%）为共显性标记，220 个（85.27%）为非同义 SNP，257 个（99.6%）为新开发的标记。此外，53 个标记用于 34 个白菜资源的基因型分析。聚类分析根据结球类型将这 34 个材料分为 3 个聚类。本研究中的数百万个 SNP 位点、大量的 KASP 标记资源以及新开发的 KASP 标记，可能会促进白菜的进一步遗传和分子育种研究。
