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A new SNP genotyping technology Target SNP-seq and its application in genetic analysis of cucumber varieties.

ABSTRACT: To facilitate the utility of SNP-based genotyping, we developed a new method called target SNP-seq which combines the advantages of multiplex PCR amplification and high throughput sequencing. Compared with KASP, Microarrays, GBS and other SNP genotyping methods, target SNP-seq is flexible both in SNPs and samples, yields high accuracy, especially when genotyping genome wide perfect SNPs with high polymorphism and conserved flanking sequences, and is cost-effective, requiring 3 days and $7 for per DNA sample to genotype hundreds of SNP loci. The present study established a DNA fingerprint of 261 cucumber varieties by target SNP-seq with 163 perfect SNPs from 4,612,350 SNPs based on 182 cucumber resequencing datasets. Four distinct subpopulations were found in 261 Chinese cucumber varieties: the north China type, the south China type, the Europe type, and the Xishuangbanna type. The north China type and Xishuangbanna type harbored lower genetic diversity, indicating greater risk of genetic erosion in these two subpopulations. Furthermore, a core set of 24 SNPs was able to distinguish 99% of the 261 cucumber varieties. 29 core cucumber backbone varieties in China were identified. Therefore, target SNP-seq provides a new way to screen out core SNP loci from the whole genome for DNA fingerprinting of crop varieties. The high efficiency and low cost of target SNP-seq is more competitive than the current SNP genotyping methods, and it has excellent application prospects in genetic research, as well as in promoting plant breeding processes in the near future.

SUBMITTER: Zhang J

PROVIDER: S-EPMC7101363 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe widely cultivated pepper (Capsicum spp.) is one of the most diverse vegetables; however, little research has focused on characterizing the genetic diversity and relatedness of commercial varieties grown in China. In this study, a panel of 92 perfect single-nucleotide polymorphisms (SNPs) was identified using re-sequencing data from 35 different C. annuum lines. Based on this panel, a Target SNP-seq genotyping method was designed, which combined multiplex amplification of perfect SNPs with Illumina sequencing, to detect polymorphisms across 271 commercial pepper varieties.ResultsThe perfect SNPs panel had a high discriminating capacity due to the average value of polymorphism information content, observed heterozygosity, expected heterozygosity, and minor allele frequency, which were 0.31, 0.28, 0.4, and 0.31, respectively. Notably, the studied pepper varieties were morphologically categorized based on fruit shape as blocky-, long horn-, short horn-, and linear-fruited. The long horn-fruited population exhibited the most genetic diversity followed by the short horn-, linear-, and blocky-fruited populations. A set of 35 core SNPs were then used as kompetitive allele-specific PCR (KASPar) markers, another robust genotyping technique for variety identification. Analysis of genetic relatedness using principal component analysis and phylogenetic tree construction indicated that the four fruit shape populations clustered separately with limited overlaps. Based on STRUCTURE clustering, it was possible to divide the varieties into five subpopulations, which correlated with fruit shape. Further, the subpopulations were statistically different according to a randomization test and Fst statistics. Nine loci, located on chromosomes 1, 2, 3, 4, 6, and 12, were identified to be significantly associated with the fruit shape index (p < 0.0001).ConclusionsTarget SNP-seq developed in this study appears as an efficient power tool to detect the genetic diversity, population relatedness and molecular breeding in pepper. Moreover, this study demonstrates that the genetic structure of Chinese pepper varieties is significantly influenced by breeding programs focused on fruit shape.

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A new SNP genotyping technology Target SNP-seq and its application in genetic analysis of cucumber varieties.

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