update ICELL8 5' scRNA TSO adapter and non-UMI technology (25c5afc0) · Commits · github_fork / Universc

launch_universc.sh

+30 −73

Original line number	Diff line number	Diff line
		@@ -645,7 +645,7 @@ elif [[ "$technology" == "icell8-non-umi" ]] \|\| [[ "$technology" == "icell8-nonu
		nonUMI=true
		elif [[ "$technology" == "icell8-five-prime" ]] \|\| [[ "$technology" == "icell8fiveprime" ]] \|\| [[ "$technology" == "icell8-5p" ]] \|\| [[ "$technology" == "icell85p" ]]; then
		technology="icell8-5-prime"
		nonUMI=false
		nonUMI=true
		if [[ -z ${chemistry} ]] \|\| [[ ${chemistry} == "SC3Pv"* ]]; then
		if [[ $verbose ]]; then
		echo "setting chemistry for 5' scRNA"
		@@ -815,7 +815,11 @@ elif [[ "$technology" == "icell8" ]]; then
		minlength=11
		elif [[ "$technology" == "icell8-5-prime" ]]; then
		barcodelength=10
		if [[ $nonUMI ]]; then
		umilength=0
		else
		umilength=8
		fi
		minlength=10
		elif [[ "$technology" == "indrop-v1" ]] \|\| [[ "$technology" == "indrop-v2" ]]; then
		barcodelength=19
		@@ -2785,6 +2789,7 @@ else
		fi

		if [[ $nonUMI ]]; then
		if [[ $chemistry == "SC3P"* ]] && [[ "$technology" == "icell8" ]]; then
		#remove inflated umi (to replace with mock and count as reads)
		sed -E '
		/^(.{11})(.{14})(.*)/ {
		@@ -2794,8 +2799,9 @@ else
		s/^(.{11})(.{14})(.*)/\1\3/g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		fi

		# add mock UMI (count reads instead of UMI) barcodelength=16, umi_default=10
		# add mock UMI (count reads instead of UMI) barcodelength=10 or 11, umi_default=10 <- default for "icell8-5-prime" and "icell8-full-length"
		perl sub/AddMockUMI.pl --fastq=${convR1} --out_dir $crIN --head_length=$barcodelength --umi_length=$umi_default
		umilength=$umi_default
		umiadjust=0
		@@ -2812,88 +2818,39 @@ else
		if [[ $chemistry == "SC5P"* ]] \|\| [[ $chemistry == "five"* ]]; then
		#remove TSO adapters (from ends)
		sed -E '
		/^TTACTATGCCGCTGGTGGCTCTAGATGTGAGAAAGGG/ {
		s/^TTACTATGCCGCTGGTGGCTCTAGATGTGAGAAAGGG/GGG/g
		n
		n
		s/^(.{34})(.{3})/\2//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^AGATCGGAAGAGCGTCGTGTAGGG/ {
		s/^AGATCGGAAGAGCGTCGTGTAGGG/GGG/g
		n
		n
		s/^(.{21})(.{3})/\2//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^GCGTCGTGTAGGG/ {
		s/^GCGTCGTGTAGGG/GGG/g
		/^(.{10})(.{10})GCGTCGTGTAGGG/ {
		s/^(.{10})(.{10})GCGTCGTGTAGGG/\1\2GGG/g
		n
		n
		s/^(.{10})(.{3})/\2//g
		s/^(.{10})(.{10})(.{10})(.{3})/\1\2\4//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^TGTGAGAAAGGG/ {
		s/^TGTGAGAAAGGG/GGG/g
		/^(.{10})(.{10})TGTGAGAAAGGG/ {
		s/^(.{10})(.{10})TGTGAGAAAGGG/\1\2GGG/g
		n
		n
		s/(.{9})(.{3})/\2//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		#remove TSO adapters (from after cell barcode) #<- confirm tag sequence with Takara reps (appears to be CB-UMI-GGG)
		sed -E '
		/^(.{11})(.{14})TTACTATGCCGCTGGTGGCTCTAGATGTGAGAAAGGG/ {
		s/^(.{11})(.{14})TTACTATGCCGCTGGTGGCTCTAGATGTGAGAAAGGG/\1\2GGG/g
		n
		n
		s/^(.{11})(.{14})(.{34})(.{3})/\1\2\4//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^(.{11})(.{14})AGATCGGAAGAGCGTCGTGTAGGG/ {
		s/^(.{11})(.{14})AGATCGGAAGAGCGTCGTGTAGGG/\1\2GGG/g
		n
		n
		s/^(.{11})(.{14})(.{21})(.{3})/\1\2\4//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^(.{11})(.{14})GCGTCGTGTAGGG/ {
		s/^(.{11})(.{14})GCGTCGTGTAGGG/\1\2GGG/g
		n
		n
		s/^(.{11})(.{14})(.{10})(.{3})/\1\2\4//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		sed -E '
		/^(.{11})(.{14})TGTGAGAAAGGG/ {
		s/^(.{11})(.{14})TGTGAGAAAGGG/\1\2GGG/g
		n
		n
		s/(.{11})(.{14})(.{9})(.{3})/\1\2\4//g
		s/(.{10})(.{10})(.{9})(.{3})/\1\2\4//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		#convert TSO to expected length for 10x 5' (TSS in R1 from base 39)
		echo " handling $convFile ..."
		tsoS="TTTCTTATATGGG" #<- confirm tag sequence with Takara reps
		tsoQ="IIIIIIIIIIIII"
		#Add 10x TSO characters to the end of the sequence (remove 'CCGGG' linker)
		cmd=$(echo 'sed -E "2~4s/(.{'$barcodelength'})(.{'${umilength}'})(.{5})/\1\2'$tsoS'/" '$convFile' > '${crIN}'/.temp') #<- confirm tag sequence (GGG) with Takara reps
		#Add 10x TSO characters to the end of the sequence (remove 'GGG' linker)
		cmd=$(echo 'sed -E "2~4s/(.{'$barcodelength'})(.{'${umilength}'})(.{3})/\1\2'$tsoS'/" '$convFile' > '${crIN}'/.temp') #<- confirm tag sequence (GGG) with Takara reps
		if [[ $verbose ]]; then
		echo technology $technology
		echo barcode: $barcodelength
		echo umi: $umilength
		echo $cmd
		fi
		# run command with barcode and umi length, e.g.,: sed -E "2~4s/(.{10})(.{8})(.{5})(.*)/\1\2$tsoS\4/" $convFile > ${crIN}/.temp
		# run command with barcode and umi length, e.g.,: sed -E "2~4s/(.{10})(.{10})(.{3})(.*)/\1\2$tsoS\4/" $convFile > ${crIN}/.temp
		eval $cmd
		mv ${crIN}/.temp $convFile
		#Add n characters to the end of the quality
		cmd=$(echo 'sed -E "4~4s/(.{'$barcodelength'})(.{'${umilength}'})(.{5})/\1\2'$tsoQ'/" '$convFile' > '${crIN}'/.temp')
		# run command with barcode and umi length, e.g.,: sed -E "4~4s/(.{10})(.{8})(.{5})(.*)/\1\2$tsoQ\4/" $convFile > ${crIN}/.temp
		cmd=$(echo 'sed -E "4~4s/(.{'$barcodelength'})(.{'${umilength}'})(.{3})/\1\2'$tsoQ'/" '$convFile' > '${crIN}'/.temp')
		# run command with barcode and umi length, e.g.,: sed -E "4~4s/(.{10})(.{10})(.{3})(.*)/\1\2$tsoQ\4/" $convFile > ${crIN}/.temp
		eval $cmd
		mv ${crIN}/.temp $convFile
		fi

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