title: Development of intron targeting (IT) markers specific for chromosome arm 4VS of Haynaldia villosa by chromosome sorting and next-generation sequencing.
abstract: Haynaldia villosa (L.) Schur (syn. Dasypyrum villosum L. Candargy, 2n = 14, genome VV) is the tertiary gene pool of wheat, and thus a potential resource of genes for wheat improvement. Among other, wheat yellow mosaic (WYM) resistance gene Wss1 and a take-all resistance gene were identified on the short arm of chromosome 4 V (4VS) of H. villosa. We had obtained introgressions on 4VS chromosome arm, with the objective of utilizing the target genes. However, monitoring these introgressions has been a daunting task because of inadequate knowledge as to H.villosa genome, as reflected by the lack of specific markers.
  
  
  
  This study aims to develop 4VS-specific markers by combination of chromosome sorting and next-generation sequencing. The short arm of chromosome 4VS of H.villosa was flow-sorted using a FACSVantage SE flow cytometer and sorter, and then sequenced by Illumina sequencing. The sequence of H. villosa 4VS was assembled by the software Hecate, and then was compared with the sequence assemblies of wheat chromosome arms 4AL, 4BS and 4DS and Ae. tauschii 4DS, with the objectives of identifying exon-exon junctions and localizing introns on chromosome 4VS of H. villosa. The intron length polymorphisms suitable for designing H. villosa primers were evaluated with criteria. Consequently, we designed a total of 359 intron targeting (IT) markers, among which 232 (64.62%) markers were specific for tracing the 4VS chromatin in the wheat background.
  
  
  
  The combination of chromosome sorting and next-generation sequencing to develop specific IT markers for 4VS of H. villosa has high success rate and specificity, thus being applicable for the development of chromosome-specific markers for alien chromatin in wheat breeding.
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Keli
authors.initials: K
authors.lastname: Dai
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Xiao
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Chunxia
authors.initials: C
authors.lastname: Yuan
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Renhui
authors.initials: R
authors.lastname: Zhao
authors.affiliation: Institute of Experimental Botany, Centre of the Haná Region for Biotechnological and Agricultural Research, Šlechtitelů 31, CZ-783671, Olomouc, Czech Republic.
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Doležel
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Aizhong
authors.initials: A
authors.lastname: Cao
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Peidu
authors.initials: P
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China.
authors.forename: Shouzhong
authors.initials: S
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Crop Genetics and Germplasm Enhancement, Cytogenetics Institute, Nanjing Agricultural University/JCIC-MCP, Nanjing, 210095, China. xiuew@njau.edu.cn.
authors.forename: Xiue
authors.identifier: 0000-0002-6312-3417
authors.initials: X
authors.lastname: Wang
mesh_terms: D001947:Breeding / Q000379:methods*; D032461:Chromosomes, Plant / Q000235:genetics*; D005819:Genetic Markers / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D007438:Introns / Q000235:genetics*; D006109:Poaceae / Q000235:genetics*; D014908:Triticum / Q000235:genetics
chemical_list: D005819:Genetic Markers
keywords: Haynaldia villosa; Intron polymorphism; Molecular marker; Triticum aestivum