Development and validation of allele-specific SNP/indel markers for eight yield-enhancing genes using whole-genome sequencing strategy to increase yield potential of rice, Oryza sativa L.

Abstract

BackgroundRice is one of the major staple foods in the world, especially in the developing countries of Asia. Its consumption as a dietary source is also increasing in Africa. To meet the demand for rice to feed the increasing human population, increasing rice yield is essential. Improving the genetic yield potential of rice is one ideal solution. It is imperative to introduce the identified yield-enhancing gene(s) into modern rice cultivars for the rapid improvement of yield potential through marker-assisted breeding.ResultsWe report the development of PCR-gel-based markers for eight yield-related functional genes (Gn1a, OsSPL14, SCM2, Ghd7, DEP1, SPIKE, GS5, and TGW6) to introduce yield-positive alleles from the donor lines. Six rice cultivars, including three each of donor and recipient lines, respectively, were sequenced by next-generation whole-genome sequencing to detect DNA polymorphisms between the genotypes. Additionally, PCR products containing functional nucleotide polymorphism (FNP) or putative FNPs for yield-related genes were sequenced. DNA polymorphisms discriminating yield-positive alleles and non-target alleles for each gene were selected through sequence analysis and the allele-specific PCR-gel-based markers were developed. The markers were validated with our intermediate breeding lines produced from crosses between the donors and 12 elite indica rice cultivars as recipients. Automated capillary electrophoresis was tested and fluorescence-labeled SNP genotyping markers (Fluidigm SNP genotyping platform) for Gn1a, OsSPL14, Ghd7, GS5, and GS3 genes were developed for high-throughput genotyping.ConclusionsThe SNP/indel markers linked to yield related genes functioned properly in our marker-assisted breeding program with identified high yield potential lines. These markers can be utilized in local favorite rice cultivars for yield enhancement. The marker designing strategy using both next generation sequencing and Sanger sequencing methods can be used for suitable marker development of other genes associated with useful agronomic traits.Electronic supplementary materialThe online version of this article (doi:10.1186/s12284-016-0084-7) contains supplementary material, which is available to authorized users.

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“…In contrast to our results, Lee et al [ 42 ] could achieve three type of alleles by employing the markers developed from the promoter region of GS5 gene which showed significant association with all the grain size traits. This result indicates that the RM574 is not efficient enough to determine the allelic distribution of GS5 gene for rice grain size in comparison to the genic variations in the promoter sequence [ 42 , 46 ].…”

GBS-based single dosage markers for linkage and QTL mapping allow gene mining for yield-related traits in sugarcane

“…Currently, high-throughput next-generation sequencing (NGS) technologies have provided new opportunities for discovering molecular markers, especially single nucleotide polymorphisms (SNPs), at the genome-wide level [41–43]. Some of these techniques, e.g.…”

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“…e sequences, which included both upstream and downstream regions of the target SNPs (Supplement 1), were sent to Fluidigm (http://Assay_Design_Group@fluidigm.com), and the Fluidigm SNP genotyping markers which was composed of a specific target amplification (STA) primer, a locus-specific (LS) primer, and two allele-specific primers were designed. Genotyping was performed following the Fluidigm SNP genotyping instructions by the IRRI genotyping service laboratory (http://gsl@irri.org) as introduced by Kim et al [31]. Briefly, the target region was amplified with the STA and LS primers under a thermal cycler.…”