SNP detection in Pinus pinaster transcriptome and association with resistance to pinewood nematode
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Date
Authors
Modesto, Ines
Inacio, Vera
Novikova, Polina
Carrasquinho, Isabel
Van de Peer, Yves
Miguel, Celia
Journal Title
Journal ISSN
Volume Title
Publisher
MDPI
Abstract
Pinewood nematode (PWN, Bursaphelenchus xylophilus) is the causal agent of pine wilt disease (PWD), which severely affects Pinus pinaster stands in southwestern Europe. Despite the high susceptibility of P. pinaster, individuals of selected half-sib families have shown genetic variability in survival after PWN inoculation, indicating that breeding for resistance can be a valuable strategy to control PWD. In this work, RNA-seq data from susceptible and resistant plants inoculated with PWN were used for SNP discovery and analysis. A total of 186,506 SNPs were identified, of which 31 were highly differentiated between resistant and susceptible plants, including SNPs in genes involved in cell wall lignification, a process previously linked to PWN resistance. Fifteen of these SNPs were selected for validation through Sanger sequencing and 14 were validated. To evaluate SNP-phenotype associations, 40 half-sib plants were genotyped for six validated SNPs. Associations with phenotype after PWN inoculation were found for two SNPs in two different genes (MEE12 and PCMP-E91), as well as two haplotypes of HIPP41, although significance was not maintained following Bonferroni correction. SNPs here detected may be useful for the development of molecular markers for PWD resistance and should be further investigated in future association studies.
Description
SUPPLEMENTARY MATERIALS : Supplementary Figure S1. Type of SNPs identified
in P. pinaster RNA-seq analysis; Supplementary Figure S2. Boxplots of the height and diameter at
the base of the stem of inoculated plants (half-sib family 440) and t-test results for the comparison
of these parameters’ means between susceptible and resistant plants; Supplementary Figure S3.
Association analysis of the SNPs in the six sequenced gene fragments under different genetic models
with resistance to PWN; Supplementary Figure S4. Genotypes distribution for SNPs associated
with phenotype; Supplementary Table S1. Summary of PCR conditions and sequencing results of
the 26 SNPs selected for validation; Supplementary Table S2. Summary of mapping statistic per
sample and per sequencing lane; Supplementary Table S3. SNPs detected in P. pinaster RNA-seq
data; Supplementary Table S4. Details and functional annotation of the SNPs with an Fst ≥ 0.80;
Supplementary Table S5. Allele frequencies and Hardy–Weinberg Equilibrium significance values
calculated by SNPassoc; Supplementary Table S6. Non-significant results of the haplotype association
analysis obtained with SNPassoc. All analyses were performed using a logistics regression model.
DATA AVAILABILITY STATEMENT : The sequencing data presented in this study are openly available in European Nucleotide Archive (ENA) at EMBL EBI under accession number PRJEB51636.
DATA AVAILABILITY STATEMENT : The sequencing data presented in this study are openly available in European Nucleotide Archive (ENA) at EMBL EBI under accession number PRJEB51636.
Keywords
Maritime pine, RNA-seq, Molecular markers, Pinewood nematode (Bursaphelenchus xylophilus), Pine wilt disease (PWD), Single nucleotide polymorphism (SNP)
Sustainable Development Goals
Citation
Modesto, I.; Inácio, V.;
Novikova, P.; Carrasquinho, I.; Van
de Peer, Y.; Miguel, C.M. SNP
Detection in Pinus pinaster
Transcriptome and Association with
Resistance to Pinewood Nematode.
Forests 2022, 13, 946. https://doi.org/10.3390/f13060946.