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PANZ_SV_Annotation.sh
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PANZ_SV_Annotation.sh
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#!/usr/bin/env bash
# set -o xtrace
# set -o errexit
set -o nounset
set -o pipefail
# >>>>>>>>>>>>>>>>>>>>>>>> Common functions >>>>>>>>>>>>>>>>>>>>>>>>
gst_log () {
local info=$1
echo -e "\033[36m[$(date +'%y-%m-%d %H:%M')]\033[0m $info" >&2
}
gst_err () {
local info=$1
echo -e "\033[31m\033[7m[ERROR]\033[0m --> $info" >&2
}
gst_warn () {
local info=$1
echo -e "\033[35m[WARNING]\033[0m --> $info" >&2
}
check_files_exists(){
local num_related_file=1
local related_file=""
for related_file in "$@"
do
if [[ ! -s "$related_file" ]]; then
echo -e "\033[31m\033[7m[ERROR]\033[0m --> No file: $related_file $" >&2
let num_related_file++
fi
done
[ "$num_related_file" -ne 1 ] && exit 1
}
check_sftw_path(){
local num_tp_program=1
local tp_program=""
for tp_program in "$@"
do
if ! which $tp_program >/dev/null 2>&1 ; then
echo -e "\033[31m\033[7m[ERROR]\033[0m --> Program not in PATH: $tp_program $" >&2
let num_tp_program++
fi
done
[ "$num_tp_program" -ne 1 ] && exit 1
}
check_var_empty () {
local var_cc=1
local var_name=""
local var=""
for var_name in "$@"; do
var=$(eval echo "$"$var_name)
case ${var} in
'')
echo -e "\033[31m\033[7m[ERROR]\033[0m --> $var_name is empty: '$var' " >&2
let var_cc++ ;;
*) ;;
esac >&2
done
[ "$var_cc" -ne 1 ] && exit 1
}
check_var_numeric () {
local var_cc=1
local var_name=""
local var=""
for var_name in "$@"; do
var=$(eval echo "$"$var_name)
# add ${var#prefix} substitution to trim sign
case ${var#[-+]} in
'')
echo -e "\033[31m\033[7m[ERROR]\033[0m --> $var_name is empty: '$var' " >&2
let var_cc++ ;;
*.*.*)
echo -e "\033[31m\033[7m[ERROR]\033[0m --> $var_name has more than one decimal point: '$var' " >&2
let var_cc++ ;;
*[!0-9.]*)
echo -e "\033[31m\033[7m[ERROR]\033[0m --> $var_name has a non-digit somewhere in it: '$var' " >&2
let var_cc++ ;;
*) ;;
esac >&2
done
[ "$var_cc" -ne 1 ] && exit 1
}
check_suffix () {
check_suffix_file=$( basename $1 )
check_suffix=$2
# add x incase file has no suffix
if [[ "${check_suffix_file##*.}"x != "$check_suffix"x ]];then
echo "[ERROR] --> $check_suffix_file should have suffix: '$check_suffix'." >&2
exit 1
fi
}
export -f gst_log gst_warn gst_err check_var_empty check_var_numeric check_sftw_path check_suffix
# <<<<<<<<<<<<<<<<<<<<<<<< Common functions <<<<<<<<<<<<<<<<<<<<<<<<
usage="
------------------------------------------------------------
PANZ annotate SV with TE and Gene gff
------------------------------------------------------------
Dependence in PATH: parallel, RepeatMasker, bedtools, bcftools
------------------------------------------------------------
USAGE:
bash $(basename $0) [OPTIIONS]
OPTIONS:
1. SV.vcf
2. gene.gff
3. TE.bed: chr start end ID class strand
4. TE.fa: NR TE sequences
5. Flanking_length(bp)
6. Threads
7. output_name
------------------------------------------------------------
Author: Songtao Gui
E-mail: [email protected]
"
if [[ $# -ne 7 ]]; then
echo "$usage" >&2
exit 1
fi
vcf=$1
gene_gff=$2
te_bed=$3
te_fa=$4
export flank=$5
threads=$6
export output=$7
overlap_TE=0.5
check_sftw_path parallel RepeatMasker bedtools bcftools
check_files_exists $vcf $gene_gff $te_bed $te_fa
check_var_numeric flank threads
check_var_empty output
# ? dealing with TEs
# ? Generate Non-Ins.bed and Ins.fa
gst_log "Dealing with TEs ..."
if [ -s "${output}/01_TE_Anno.tsv" ];then
gst_warn "Skip running, result file exists."
else
gst_log "Generate Non-Ins.bed and Ins.fa ..."
if [ -s "${output}/Non_INS.bed" -a -s "${output}/INS.fa" ];then
gst_warn "Skip running, result file exists."
else
bcftools view --threads $threads $vcf | parallel --pipe -q -k -j $threads perl -F"\t" -lane '
BEGIN{$,="\t";}
/^#/ && next;
if(/SVTYPE=INS/){
$type="INS";
$out=join("\t",@F[2,4]);
# ! RepeatMasker limits the Fasta max id length to 50
print "$type,$out" if ++$h{$F[2]}==1 and length($F[2]) < 50;
}else{
$type="Non_INS";
($id,$c1,$p1,$c2,$p2)=split("#",$F[2]);
if($c1 eq $c2){
print "$type,$c1",$p1-1,$p2,$F[2] if ++$h{$F[2]}==1;
}else{
print "$type,$c1",$p1-1,$p1,$F[2] if ++$h{$F[2]}==1;
print "$type,$c2",$p2-1,$p2,$F[2] if ++$h{$F[2]}==1;
}
}
' | csvtk split -H -f 1 -o ${output}
if [ $? -ne 0 ];then gst_err "get ins/nonins bed fa failed: Non-zero exit";rm -rf ${output}; exit 1;fi
# ? format bed and fa --> ${output}/Non_INS_Anno.tsv
check_files_exists ${output}/stdin-INS.csv ${output}/stdin-Non_INS.csv
csvtk cut -j $threads -f 2 ${output}/stdin-INS.csv | seqkit tab2fx -j $threads -o ${output}/INS.fa &&\
csvtk cut -j $threads -f 2 ${output}/stdin-Non_INS.csv -o ${output}/Non_INS.bed &&\
rm -f ${output}/stdin-INS.csv ${output}/stdin-Non_INS.csv
if [ $? -ne 0 ];then gst_err "format bed and fa failed: Non-zero exit"; exit 1;fi
check_files_exists ${output}/Non_INS.bed ${output}/INS.fa
seqkit fx2tab -n -j $threads -i ${output}/INS.fa | parallel --pipe -q -k -j $threads perl -F"#" -lane '
$,="\t";
$F[-1]=~s/\s+$//;
print $F[1],$F[2]-1,$F[4],join("#",@F);
' > ${output}/INS.bed
fi
gst_log "Check Non_INS SV that >= $overlap_TE overlapped with TEs ..."
if [ ! -s "${output}/Non_INS_Anno.tsv" ];then
bedtools intersect -e -f $overlap_TE -F $overlap_TE -a ${output}/Non_INS.bed -b $te_bed -wo |\
parallel --pipe -j $threads -q -k perl -lane 'print "$F[3]\t$F[7]#OVLP_$F[10]" if $F[10]' |\
csvtk collapse -j $threads -tTH -f 1 -v 2 -s "," -o ${output}/Non_INS_Anno.tsv
if [ $? -ne 0 ];then gst_err "Annotate Non_INS failed: Non-zero exit";rm -f ${output}/Non_INS_Anno.tsv; exit 1;fi
else
gst_warn "Skip running, result file exists."
fi
# ? RM INS.fa with cutoff 250 --> ${output}/INS_Anno.tsv
gst_log "Identify TE INS using RepeatMasker against $te_fa ..."
if [ ! -s "${output}/INS_Anno.tsv" ];then
RepeatMasker -cutoff 250 -nolow -no_is -norna -parallel $threads -lib $te_fa -dir ${output}/INS_RM ${output}/INS.fa
if [ $? -ne 0 ];then gst_err "RepeatMasker for INS.fa failed: Non-zero exit";rm -rf ${output}/INS_RM; exit 1;fi
# ? Parsing RM out
check_files_exists ${output}/INS_RM/INS.fa.out
sed '1,3d' ${output}/INS_RM/INS.fa.out | parallel --pipe -j $threads -q -k perl -lane 'print "$F[4]\t$F[9]#SW_$F[0]" if ++$h{$F[4]}==1' > ${output}/INS_Anno.tsv
if [ $? -ne 0 ];then gst_err "Parse RM out failed: Non-zero exit"; rm -f ${output}/INS_Anno.tsv; exit 1;fi
else
gst_warn "Skip running, result file exists."
fi
gst_log "merge TE annotation"
cat ${output}/Non_INS_Anno.tsv ${output}/INS_Anno.tsv | csvtk sort -j $threads -tTH -k 1 -o ${output}/01_TE_Anno.tsv
fi
gst_log "Done for TE annotation!
"
gst_log "Dealing with Genes ..."
# ? get All SV Bed
if [ ! -s "${output}/ALL_SV_flanking.bed" ];then
check_files_exists ${output}/INS.bed ${output}/Non_INS.bed
cat ${output}/INS.bed ${output}/Non_INS.bed | parallel --pipe -j $threads -q -k perl -F"\t" -lane '
$fa=$F[1]-$ENV{flank}; $fa=0 if $fa < 0;
$fb=$F[2]+$ENV{flank};
$,="\t";
print $F[0],$fa,$fb,join("|",@F);
' | csvtk sort -tTH -k 1 -k 2:n -k 3:n -j $threads -o ${output}/ALL_SV_flanking.bed
if [ $? -ne 0 ];then gst_err "get All flank bed failed: Non-zero exit";rm -f ${output}/ALL_SV_flanking.bed; exit 1;fi
fi
# ? filter gene gff to include only col3 =~ /RNA|CDS|UTR/ --> /RNA/
gst_log "Formatting annotation gff ..."
if [ ! -s "${output}/fmtted_gene.bed" ];then
cat $gene_gff | parallel --pipe -q -k -j $threads perl -F"\t" -lane '
next unless $F[2]=~/RNA/;
$id="NA";
if($F[-1]=~/ID=(.*?);/){
$id=$1;
}elsif($F[-1]=~/Parent=([^\s;]+)/){
$id="$1"."_$F[2]";
}else{
next;
}
$,="\t";
#$num=++$h{"$id,$F[6]"};
#print @F[0,3,4],"$id\#$num",@F[2,6];
print @F[0,3,4],$id,@F[2,6];
' > ${output}/fmtted_gene.bed
if [ $? -ne 0 ];then gst_err "gt fmtted_gene.bed failed: Non-zero exit"; rm -f ${output}/fmtted_gene.bed; exit 1;fi
fi
# ? overlap with genes
# USAGE:0 <pipe with bedtools intersect output>
# 0 1 2 3 4 5 6 7 8 9 10
# 1 37099 326679 PZ00001aSV00000102DEL 1 44351 44947 CDS:Zm00001d027230_P001_1 CDS + 596
fmt_gene_ovlp () {
# format gene ovlp result from intersectBed to final results
local line=""
while read line
do
echo "$line" | perl -F"\t" -lane '
$,="\t";
($c,$s,$e,$id)=split(/\|/,$F[3]);
print $c,$s,$e,$id,@F[4..$#F-1];
' |\
bedtools overlap -i stdin -cols 2,3,6,7 |\
perl -F"\t" -lane '
$,="\t";
$ovlp=abs($F[-1]);
$tag="";
if($F[-1] < 0){
# ? no ovlp
if($F[2] < $F[5]){
# ? upstream
$tag="Upstream";
}else{
# ? downstream
$tag="Downstream";
}
}else{
# ? ovlp
$tag="Overlap";
$tag="embed" if $F[1] >= $F[5] and $F[2] <= $F[6];
$tag="contain" if $F[1] < $F[5] and $F[2] > $F[6];
}
print $F[3],"$F[7]#$tag";
'
done
}
export -f fmt_gene_ovlp
gst_log "Get overlapped gene features with flanking $flank bp ..."
if [ ! -s "${output}/02_gene_anno.tsv" ];then
bedtools intersect -a ${output}/ALL_SV_flanking.bed -b ${output}/fmtted_gene.bed -wo |\
parallel --pipe -j $threads -q -k fmt_gene_ovlp |\
csvtk collapse -j $threads -tTH -f 1 -v 2 -s "," -o ${output}/02_gene_anno.tsv
else
gst_warn "Skip running, result file exists."
fi
gst_log "Done Gene annotate!
"
gst_log "Adding annotations to VCF file ..."
if [ ! -s "${output}/00_Annotated_variants.vcf.gz" ];then
gst_log "Formatting final vcf ..."
bcftools view --threads $threads $vcf | perl -F"\t" -lane '
BEGIN{
$tefile="$ENV{output}/01_TE_Anno.tsv";
open(TE,"$tefile") or die("Cannot open file: $tefile");
while(<TE>){
chomp;
# $hash{$_}=1;
($var1,$var2)=split(/\t/,$_);
$TE_hash{$var1}=$var2;
}
close TE;
$genefile="$ENV{output}/02_gene_anno.tsv";
open(GENE,"$genefile") or die("Cannot open file: $genefile");
while(<GENE>){
chomp;
# $hash{$_}=1;
($var1,$var2)=split(/\t/,$_);
$GENE_hash{$var1}=$var2;
}
close GENE;
$,="\t";
}
/^##/ && print && next;
if(/^#CHROM/){
print "##INFO=<ID=TE_ANNO,Number=1,Type=String,Description=\"Annotation of overlapped TEs\">\n##INFO=<ID=GENE_ANNO,Number=1,Type=String,Description=\"Annotation of Genes overlapped with flanking $ENV{flank} bp\">";
print $_;
next;
}
if($TE_hash{$F[2]}){
$F[7]=$F[7].";TE_ANNO=$TE_hash{$F[2]}";
}
if($GENE_hash{$F[2]}){
$F[7]=$F[7].";GENE_ANNO=$GENE_hash{$F[2]}";
}
print @F if ++$h{$F[2]}==1;
' | bcftools view --threads $threads -O z -o ${output}/00_Annotated_variants.vcf.gz
if [ $? -ne 0 ];then gst_err "get annotated vcf failed: Non-zero exit";rm -f ${output}/00_Annotated_variants.vcf.gz; exit 1;fi
fi
if [ ! -s "${output}/00_Annotated_variants.vcf.gz.tbi" ];then
gst_log "Indexing ..."
bcftools index --threads $threads -t ${output}/00_Annotated_variants.vcf.gz
if [ $? -ne 0 ];then gst_err "index failed: Non-zero exit"; exit 1;fi
fi
gst_log "Yes!! All Done !!"