Abstract
The use of molecular markers in plant breeding has become a routine practice, but the cost per accession can be a hindrance to the routine use of Quantitative Trait Loci (QTL) identification in breeding programs. In this study, we demonstrate the use of targeted re-sequencing as a proof of concept of a cost-effective approach to retrieve highly informative allele information, as well as develop a bioinformatics strategy to capture the genome-specific information of a polyploid species. SNPs were identified from alignment of raw transcriptome reads (2 × 50 bp) to a synthetic tetraploid genome using BWA followed by a GATK pipeline. Regions containing high polymorphic SNPs in both A genome and B genomes were selected as targets for the resequencing study. Targets were amplified using multiplex PCR followed by sequencing on an Illumina HiSeq. Eighty-one percent of the SNP calls in diploids and 68% of the SNP calls in tetraploids were confirmed. These results were also confirmed by KASP validation. Based on this study, we find that targeted resequencing technologies have potential for obtaining maximum allele information in allopolyploids at reduced cost.
Highlights
Marker-assisted breeding often requires a large number of samples to be genotyped, especially in early generations, for effective selection [1,2]
From the alignment with the OLin tetraploid transcriptome reference, 865 SNPs were identified with polymorphic information content greater than 0.3 and with read depth of greater than 100 (Table 3)
The current experiment was designed as a proof of concept, to test resequencing in peanut, and to determine whether it could provide a lower cost method of genotyping, at a marker density lower than high throughput methods, but sufficient for Quantitative Trait Loci (QTL) analysis and marker-assisted selection
Summary
Marker-assisted breeding often requires a large number of samples to be genotyped, especially in early generations, for effective selection [1,2]. Molecular markers such as SSRs and SNPs are the most commonly utilized types for marker-assisted selection. Sequencing) technology, high-throughput platforms have been used for SNP identification and genotyping in plants. The complexity of the polyploid plant genome has complicated the interpretation of SNP calls, and decreased validation success [3]. Challenges in genotyping polyploids have been reported before [4,5]. Bertioli et al [6] reported decreased SNP validation success from
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