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Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping

BACKGROUND: Linkage of DNA markers with phenotypic traits provides essential information to dissect clustered genes with potential phenotypic contributions in a target genome region. Pinus flexilis E. James (limber pine) is a keystone five-needle pine species in mountain-top ecosystems of North Amer...

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Autores principales: Liu, Jun-Jun, Schoettle, Anna W., Sniezko, Richard A., Sturrock, Rona N., Zamany, Arezoo, Williams, Holly, Ha, Amanda, Chan, Danelle, Danchok, Bob, Savin, Douglas P., Kegley, Angelia
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034428/
https://www.ncbi.nlm.nih.gov/pubmed/27663193
http://dx.doi.org/10.1186/s12864-016-3079-2
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author Liu, Jun-Jun
Schoettle, Anna W.
Sniezko, Richard A.
Sturrock, Rona N.
Zamany, Arezoo
Williams, Holly
Ha, Amanda
Chan, Danelle
Danchok, Bob
Savin, Douglas P.
Kegley, Angelia
author_facet Liu, Jun-Jun
Schoettle, Anna W.
Sniezko, Richard A.
Sturrock, Rona N.
Zamany, Arezoo
Williams, Holly
Ha, Amanda
Chan, Danelle
Danchok, Bob
Savin, Douglas P.
Kegley, Angelia
author_sort Liu, Jun-Jun
collection PubMed
description BACKGROUND: Linkage of DNA markers with phenotypic traits provides essential information to dissect clustered genes with potential phenotypic contributions in a target genome region. Pinus flexilis E. James (limber pine) is a keystone five-needle pine species in mountain-top ecosystems of North America. White pine blister rust (WPBR), caused by a non-native fungal pathogen Cronartium ribicola (J.C. Fisch.), has resulted in mortality in this conifer species and is still spreading through the distribution. The objective of this research was to develop P. flexilis transcriptome-wide single nucleotide polymorphism (SNP) markers using RNA-seq analysis for genetic mapping of the major gene (Cr4) that confers complete resistance to C. ribicola. RESULTS: Needle tissues of one resistant and two susceptible seedling families were subjected to RNA-seq analysis. In silico SNP markers were uncovered by mapping the RNA-seq reads back to the de novo assembled transcriptomes. A total of 110,573 in silico SNPs and 2,870 indels were identified with an average of 3.7 SNPs per Kb. These SNPs were distributed in 17,041 unigenes. Of these polymorphic P. flexilis unigenes, 6,584 were highly conserved as compared to the genome sequence of P. taeda L (loblolly pine). High-throughput genotyping arrays were designed and were used to search for Cr4-linked genic SNPs in megagametophyte populations of four maternal trees by haploid-segregation analysis. A total of 32 SNP markers in 25 genes were localized on the Cr4 linkage group (LG). Syntenic relationships of this Cr4-LG map with the model conifer species P. taeda anchored Cr4 on Pinus consensus LG8, indicating that R genes against C. ribicola have evolved independently in different five-needle pines. Functional genes close to Cr4 were annotated and their potential roles in Cr4-mediated resistance were further discussed. CONCLUSIONS: We demonstrated a very effective, low-cost strategy for developing a SNP genetic map of a phenotypic trait of interest. SNP discovery through transcriptome comparison was integrated with high-throughput genotyping of a small set of in silico SNPs. This strategy may be applied to mapping any trait in non-model plant species that have complex genomes. Whole transcriptome sequencing provides a powerful tool for SNP discovery in conifers and other species with complex genomes, for which sequencing and annotation of complex genomes is still challenging. The genic SNP map for the consensus Cr4-LG may help future molecular breeding efforts by enabling both Cr4 positional characterization and selection of this gene against WPBR. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12864-016-3079-2) contains supplementary material, which is available to authorized users.
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spelling pubmed-50344282016-09-29 Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping Liu, Jun-Jun Schoettle, Anna W. Sniezko, Richard A. Sturrock, Rona N. Zamany, Arezoo Williams, Holly Ha, Amanda Chan, Danelle Danchok, Bob Savin, Douglas P. Kegley, Angelia BMC Genomics Research Article BACKGROUND: Linkage of DNA markers with phenotypic traits provides essential information to dissect clustered genes with potential phenotypic contributions in a target genome region. Pinus flexilis E. James (limber pine) is a keystone five-needle pine species in mountain-top ecosystems of North America. White pine blister rust (WPBR), caused by a non-native fungal pathogen Cronartium ribicola (J.C. Fisch.), has resulted in mortality in this conifer species and is still spreading through the distribution. The objective of this research was to develop P. flexilis transcriptome-wide single nucleotide polymorphism (SNP) markers using RNA-seq analysis for genetic mapping of the major gene (Cr4) that confers complete resistance to C. ribicola. RESULTS: Needle tissues of one resistant and two susceptible seedling families were subjected to RNA-seq analysis. In silico SNP markers were uncovered by mapping the RNA-seq reads back to the de novo assembled transcriptomes. A total of 110,573 in silico SNPs and 2,870 indels were identified with an average of 3.7 SNPs per Kb. These SNPs were distributed in 17,041 unigenes. Of these polymorphic P. flexilis unigenes, 6,584 were highly conserved as compared to the genome sequence of P. taeda L (loblolly pine). High-throughput genotyping arrays were designed and were used to search for Cr4-linked genic SNPs in megagametophyte populations of four maternal trees by haploid-segregation analysis. A total of 32 SNP markers in 25 genes were localized on the Cr4 linkage group (LG). Syntenic relationships of this Cr4-LG map with the model conifer species P. taeda anchored Cr4 on Pinus consensus LG8, indicating that R genes against C. ribicola have evolved independently in different five-needle pines. Functional genes close to Cr4 were annotated and their potential roles in Cr4-mediated resistance were further discussed. CONCLUSIONS: We demonstrated a very effective, low-cost strategy for developing a SNP genetic map of a phenotypic trait of interest. SNP discovery through transcriptome comparison was integrated with high-throughput genotyping of a small set of in silico SNPs. This strategy may be applied to mapping any trait in non-model plant species that have complex genomes. Whole transcriptome sequencing provides a powerful tool for SNP discovery in conifers and other species with complex genomes, for which sequencing and annotation of complex genomes is still challenging. The genic SNP map for the consensus Cr4-LG may help future molecular breeding efforts by enabling both Cr4 positional characterization and selection of this gene against WPBR. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12864-016-3079-2) contains supplementary material, which is available to authorized users. BioMed Central 2016-09-23 /pmc/articles/PMC5034428/ /pubmed/27663193 http://dx.doi.org/10.1186/s12864-016-3079-2 Text en © The Author(s). 2016 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research Article
Liu, Jun-Jun
Schoettle, Anna W.
Sniezko, Richard A.
Sturrock, Rona N.
Zamany, Arezoo
Williams, Holly
Ha, Amanda
Chan, Danelle
Danchok, Bob
Savin, Douglas P.
Kegley, Angelia
Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title_full Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title_fullStr Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title_full_unstemmed Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title_short Genetic mapping of Pinus flexilis major gene (Cr4) for resistance to white pine blister rust using transcriptome-based SNP genotyping
title_sort genetic mapping of pinus flexilis major gene (cr4) for resistance to white pine blister rust using transcriptome-based snp genotyping
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034428/
https://www.ncbi.nlm.nih.gov/pubmed/27663193
http://dx.doi.org/10.1186/s12864-016-3079-2
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