Codes for the paper : "Genomic resources for Mediterranean fishes"

---

Pierre-Edouard Guerin, Stephanie Manel

Montpellier, 2017-2019

Submited to Molecular Ecology Ressources, 2019

---

Prerequisites

Softwares

• R Version 3.6.0

  • R packages: ggplot2, plyr, reshape, gridExtra

• faidx

• bedtools

• vcftools

• samtools

• htslib

• bcftools

Singularity container

See https://www.sylabs.io/docs/ for instructions to install Singularity.

Download the container

singularity pull --name snpsdata_analysis.simg shub://Grelot/reserveBenefit--snpsdata_analysis:snpsdata_analysis

Run the container

singularity run snpsdata_analysis.simg

---

Data files

We work on three species : mullus surmuletus, diplodus sargus and serranus cabrilla. Let's define the wildcard species as any of these three species.

• genome assembly .fasta

• SNPs data from radseq .vcf

Filtering SNPs

Only one randomly selected SNP was retained per locus, and a locus was retained only if present in at least 85% of individuals. Individuals with an excess coverage depth (>1,000,000x) or >30% missing data were filtered out. We kept loci with maximum observed heterozygosity=0.6.

Filtering steps (IBD paper)

1. Remove loci with inbreeding coefficient Fis > 0.5 or < -0.5
2. Keep all pairs of loci that are closer than 5000 bp
3. Keep pairs of loci with linkage desequilibrum r² > 0.8
4. Keep SNPs with a minimum minor allele frequency (MAF) of 1%
5. Remove loci that deviated significantly (p-value <0.01) from expected Hardy-Weinberg genotyping frequencies under random mating

Filtering steps (genome paper)

1. Keep all pairs of loci that are closer than 5000 bp
2. Keep pairs of loci with linkage desequilibrum r² > 0.8
3. Keep SNPs with a minimum minor allele frequency (MAF) of 1%

INPUTS:

• species.vcf: SNPs from radseq data of species
• species.sumstats.tsv: Summary statistics for each population

OUTPUTS:

• species.lmiss: number of missing individuals by locus table
• species.imiss: number of missing loci by individual table
• species.idepth: mean locus depth coverage by individual table
• species.geno.ld: linkage desequilibrum _r² table
• species.snps.fisloc_rm.vcf
• species.fisloc_rm.ld_5000.log
• species.fisloc_rm.ld_5000.recode.vcf
• species.fisloc_rm.ld_5000.r2.recode.vcf
• species.fisloc_rm.ld_5000.r2.maf001.recode.vcf
• species.fisloc_rm.ld_5000.r2.maf001.hwe.recode.vcf: final filtered snps
• speciesfiltering_count_snps_report.tsv: number of SNPs at each filtering step

cd filter_vcf
bash filter_vcf.sh

Description of SNPs onto genome

Generate tables

1. Split the genome into genome-windows of 400 Kbp.
2. Count number of SNPs located on each genome-windows.
3. Count number of reads for each SNP for each individuals.

INPUTS:

• species.fasta: genome fasta file of species
• species.vcf: SNPs from radseq data of species
• species.gff3: coordinates and related information of coding region annotation genome of species

OUTPUTS:

• speciescoverage.bed: a table with row as genome-windows of 400000bp of the genome of species with genome-coordinates (scaffold, start position, end position) and coverage (number of SNPs)
• speciesmeandepth.bed: a table with row as SNPs with genome-windows, coordinates (scaffold, start position, end position) and depth coverage (number of reads) for each SNP for each individuals
• speciescoords.snps.bed: coordinates (scaffold, position) of SNPs onto genomes
• speciescoding.snps.bed: snps located on coding region

bash snpsontothegenome/command.sh

Build the figure

Rscript snpsontothegenome/figure_cover_genome.R

Average distance between SNPs loci

INPUTS:

• speciescoords.snps.bed : coordinates (scaffold, position) of SNPs onto genomes

Rscript snpsontothegenome/average_distance_loci.R

SNPs located/not in coding regions

Simply count number of lines of the file speciescoding.snps.bed (each line is a snp located on a coding region)

SNPs located/not in mitochondrial regions

Simply count SNPs annotated as "mitochondrial" by Augustus

species	number of SNPs located in mitochondrial regions
diplodus	173
mullus	178
serran	226

Results

• distance_loci.csv : mean, median and sd distance between consecutive loci

species	mean	median	sd	max	min
diplodus	35388.9078430345	23751	34996.9143024498	459616	5000
mullus	30716.8684498214	20930	29189.8335674228	384550	5002
serran	28239.7585528699	19084	27013.2843728281	403508	733

• summary_snps.csv: number of SNPs, average distance between consecutive loci (in bp) and number of SNPs located on a coding region for each species

species	number_snps	average_distance_bp	number_coding_snps	number_mitochondrial_snps
diplodus	20074	35389	11978	173
mullus	15710	30717	10304	178
serranus	21101	28240	13107	226

More detail about coding snps location (CDS, exon, intron) in this table: count_snps_annotation.csv