QTL analysis using SNP markers developed by next-generation sequencing for identification of candidate genes controlling 4-methylthio-3-butenyl glucosinolate contents in roots of radish, Raphanus sativus L

PLoS One. 2013;8(1):e53541. doi: 10.1371/journal.pone.0053541. Epub 2013 Jan 7.

Abstract

SNP markers for QTL analysis of 4-MTB-GSL contents in radish roots were developed by determining nucleotide sequences of bulked PCR products using a next-generation sequencer. DNA fragments were amplified from two radish lines by multiplex PCR with six primer pairs, and those amplified by 2,880 primer pairs were mixed and sequenced. By assembling sequence data, 1,953 SNPs in 750 DNA fragments, 437 of which have been previously mapped in a linkage map, were identified. A linkage map of nine linkage groups was constructed with 188 markers, and five QTLs were detected in two F(2) populations, three of them accounting for more than 50% of the total phenotypic variance being repeatedly detected. In the identified QTL regions, nine SNP markers were newly produced. By synteny analysis of the QTLs regions with Arabidopsis thaliana and Brassica rapa genome sequences, three candidate genes were selected, i.e., RsMAM3 for production of aliphatic glucosinolates linked to GSL-QTL-4, RsIPMDH1 for leucine biosynthesis showing strong co-expression with glucosinolate biosynthesis genes linked to GSL-QTL-2, and RsBCAT4 for branched-chain amino acid aminotransferase linked to GSL-QTL-1. Nucleotide sequences and expression of these genes suggested their possible function in 4MTB-GSL biosynthesis in radish roots.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 2-Isopropylmalate Synthase / genetics
  • 2-Isopropylmalate Synthase / metabolism
  • Arabidopsis / genetics
  • Arabidopsis / metabolism
  • Base Sequence
  • Brassica rapa / genetics
  • Brassica rapa / metabolism
  • Chromosome Mapping
  • Chromosomes, Plant
  • DNA, Plant / genetics*
  • Expressed Sequence Tags
  • Glucosinolates / biosynthesis*
  • High-Throughput Nucleotide Sequencing
  • Malate Dehydrogenase / genetics
  • Malate Dehydrogenase / metabolism
  • Molecular Sequence Data
  • Multiplex Polymerase Chain Reaction
  • Plant Proteins / genetics*
  • Plant Proteins / metabolism
  • Plant Roots / genetics*
  • Plant Roots / metabolism
  • Polymorphism, Single Nucleotide*
  • Quantitative Trait Loci*
  • Raphanus / genetics*
  • Raphanus / metabolism
  • Synteny
  • Transaminases / genetics
  • Transaminases / metabolism

Substances

  • 4-methylthio-3-butenyl glucosinolate
  • DNA, Plant
  • Glucosinolates
  • Plant Proteins
  • Malate Dehydrogenase
  • 2-Isopropylmalate Synthase
  • Transaminases
  • branched-chain-amino-acid transaminase

Grant support

This work was supported by the Program for Promotion of Basic and Applied Research for Innovations in Bio-oriented Industry (BRAIN), Japan. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.