title: 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: BACKGROUND
  Rice 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.
  
  
  RESULTS
  We 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.
  
  
  CONCLUSIONS
  The 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.
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines.
authors.forename: Sung-Ryul
authors.initials: SR
authors.lastname: Kim
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines.
authors.forename: Joie
authors.initials: J
authors.lastname: Ramos
authors.affiliation: Bioscience and Biotechnology Center, Nagoya University, Nagoya, Japan.
authors.forename: Motoyuki
authors.initials: M
authors.lastname: Ashikari
authors.affiliation: CIAT-HarvestPlus, 303 ICRISAT Campus, Patancheru, Telungana, India.
authors.forename: Parminder S
authors.initials: PS
authors.lastname: Virk
authors.affiliation: International Center for Tropical Agriculture, A.A. 6713, Cali, Colombia.
authors.forename: Edgar A
authors.initials: EA
authors.lastname: Torres
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines.
authors.forename: Eero
authors.initials: E
authors.lastname: Nissila
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines.
authors.forename: Sherry Lou
authors.initials: SL
authors.lastname: Hechanova
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines.
authors.forename: Ramil
authors.initials: R
authors.lastname: Mauleon
authors.affiliation: Plant Breeding, Genetics, and Biotechnology Division, International Rice Research Institute, Metro Manila, Philippines. k.jena@irri.org.
authors.forename: Kshirod K
authors.initials: KK
authors.lastname: Jena
keywords: Allele-specific marker; Rice; SNP genotyping; Yield potential; Yield-enhancing genes