add inputs, documentation and subroutine for single-cell combinatorial fluidic... (4a76769f) · Commits · github_fork / Universc

README.Rmd

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Original line number	Diff line number	Diff line
		@@ -1027,6 +1027,7 @@ Mandatory arguments to long options are mandatory for short options too.
		STRT-Seq-C1 (8 bp barode, 5 bp UMI)
		STRT-Seq-2i (13 bp barcode, 6 bp UMI)
		SmartSeq2 (16 bp barcode, no UMI)
		SCIFI-Seq (27 bp barcode, 8 bp UMI

		-b, --barcodefile FILE Custom barcode list in plain text (with each line containing a barcode)

README.html

+1 −0

Original line number	Diff line number	Diff line
		@@ -496,6 +496,7 @@ Mandatory arguments to long options are mandatory for short options too.
		STRT-Seq-C1 (8 bp barode, 5 bp UMI)
		STRT-Seq-2i (13 bp barcode, 6 bp UMI)
		SmartSeq2 (16 bp barcode, no UMI)
		SCIFI-Seq (27 bp barcode, 8 bp UMI

		-b, --barcodefile FILE Custom barcode list in plain text (with each line containing a barcode)

README.md

+2 −1

Original line number	Diff line number	Diff line
		@@ -6,7 +6,7 @@ affiliations:
		index: 1
		- name: "RIKEN Center for Sustainable Resource Sciences, Suehiro-cho-1-7-22, Tsurumi Ward, Yokohama, Kanagawa 230-0045, Japan"
		index: 2
		date: "Saturday 24 April 2021"
		date: "Sunday 25 April 2021"
		output:
		prettydoc::html_pretty:
		theme: cayman
		@@ -1027,6 +1027,7 @@ Mandatory arguments to long options are mandatory for short options too.
		STRT-Seq-C1 (8 bp barode, 5 bp UMI)
		STRT-Seq-2i (13 bp barcode, 6 bp UMI)
		SmartSeq2 (16 bp barcode, no UMI)
		SCIFI-Seq (27 bp barcode, 8 bp UMI

		-b, --barcodefile FILE Custom barcode list in plain text (with each line containing a barcode)

launch_universc.sh

+67 −8

Original line number	Diff line number	Diff line
		@@ -226,6 +226,7 @@ Mandatory arguments to long options are mandatory for short options too.
		STRT-Seq-C1 (8 bp barode, 5 bp UMI)
		STRT-Seq-2i (13 bp barcode, 6 bp UMI)
		SmartSeq2 (16 bp barcode, no UMI)
		SCIFI-Seq (27 bp barcode, 8 bp UMI

		-b, --barcodefile FILE Custom barcode list in plain text (with each line containing a barcode)

		@@ -624,12 +625,14 @@ elif [[ "$technology" == "quartz-seq2-384" ]] \|\| [[ "$technology" == "quartzseq2
		technology="quartz-seq2-384"
		elif [[ "$technology" == "quartz-seq2-1536" ]] \|\| [[ "$technology" == "quartzseq2-1536" ]] \|\| [[ "$technology" == "quartz-seq2-v3.2" ]] \|\| [[ "$technology" == "quartzseq2-v3.2" ]] \|\| [[ "$technology" == "quartzseq2v3.2" ]]; then
		technology="quartz-seq2-1536"
		elif [[ "$technology" == "sciseq" ]] \|\| [[ "$technology" == "sci-seq" ]]; then
		elif [[ "$technology" == "sciseq" ]] \|\| [[ "$technology" == "sci-seq" ]] \|\| [[ "$technology" == "sci-rna-seq" ]]; then
		technology="sciseq3"
		elif [[ "$technology" == "sciseq2" ]] \|\| [[ "$technology" == "sci-seq2" ]]; then
		technology="sciseq2"
		elif [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "sci-seq3" ]]; then
		elif [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "sci-seq3" ]] \|\| [[ "$technology" == "sci-rna-seq3" ]] \|\| [[ "$technology" == "sci-rna-seq-3" ]] ; then
		technology="sciseq3"
		elif [[ "$technology" == "scifiseq" ]] \|\| [[ "$technology" == "scifi-seq" ]] \|\| [[ "$technology" == "sci-fi-seq" ]] \|\| [[ "$technology" == "scifi-rna-seq" ]]; then
		technology="scifiseq"
		elif [[ "$technology" == "scrbseq" ]] \|\| [[ "$technology" == "scrb-seq" ]] \|\| [[ "$technology" == "mcscrbseq" ]] \|\| [[ "$technology" == "mcscrb-seq" ]]; then
		technology="scrbseq"
		elif [[ "$technology" == "seqwell" ]] \|\| [[ "$technology" == "seq-well" ]]; then
		@@ -772,6 +775,10 @@ elif [[ "$technology" == "sciseq3" ]]; then
		barcodelength=40
		umilength=8
		minlength=40
		elif [[ "$technology" == "scifiseq" ]]; then
		barcodelength=27
		umilength=8
		minlength=27
		elif [[ "$technology" == "scrbseq" ]]; then
		barcodelength=6
		umilength=10
		@@ -992,7 +999,7 @@ fi
		#index 2
		if [[ $setup == "false" ]]; then
		#only check I2 for dual-indexed techniques
		if [[ "$technology" == "indrop-v3" ]] \|\| [[ "$technology" == "sciseq2" ]] \|\| [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "smartseq"* ]]; then
		if [[ "$technology" == "indrop-v3" ]] \|\| [[ "$technology" == "sciseq2" ]] \|\| [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "scifiseq" ]] \|\| [[ "$technology" == "smartseq"* ]]; then
		if [[ ${#index2[@]} -ne ${#read1[@]} ]]; then
		if [[ ${#index2[@]} -gt 0 ]]; then
		echo " Error: number of index1 files is not matching the number of index2 files"
		@@ -1519,7 +1526,7 @@ if [[ -n "$barcodefile" ]]; then
		#getting absolute path
		barcodefile=$(readlink -f $barcodefile)
		#allowing WellList from ICELL8 and other well-based techniques
		if [[ "$technology" == "bd-rhapsody" ]] \|\| [[ "$technology" == "icell8" ]] \|\| [[ "$technology" == "quartz-seq2" ]] \|\| [[ "$technology" == "microwellseq" ]] \|\| [[ "$technology" == "smartseq" ]] \|\| [[ "$technology" == "seqwell" ]] \|\| [[ "$technology" == "sciseq2" ]] \|\| [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "splitseq" ]] \|\| [[ "$technology" == "splitseq2" ]] \|\| [[ "$technology" == "custom" ]]; then
		if [[ "$technology" == "bd-rhapsody" ]] \|\| [[ "$technology" == "icell8" ]] \|\| [[ "$technology" == "quartz-seq2" ]] \|\| [[ "$technology" == "microwellseq" ]] \|\| [[ "$technology" == "smartseq" ]] \|\| [[ "$technology" == "seqwell" ]] \|\| [[ "$technology" == "sciseq2" ]] \|\| [[ "$technology" == "sciseq3" ]] \|\| [[ "$technology" == "scifiseq" ]] \|\| [[ "$technology" == "splitseq" ]] \|\| [[ "$technology" == "splitseq2" ]] \|\| [[ "$technology" == "custom" ]]; then
		seg=$'\t'
		n_col=$(awk -F'\t' '{print NF}' $barcodefile \| sort -nu \| tail -n 1)
		if [[ $n_col -eq 1 ]]; then
		@@ -1594,6 +1601,11 @@ else
		if [[ ! -f ${whitelistdir}/sciseq3_barcode.txt ]]; then
		echo " ...generating combination of I1, I2, and RT barcodes..."
		fi
		elif [[ "$technology" == "scifiseq" ]]; then
		barcodefile=${whitelistdir}/scifi-seq_barcode.txt
		if [[ ! -f ${whitelistdir}/scifi-seq_barcode.txt ]]; then
		echo " ...generating combination of I1, I2, and RT barcodes..."
		fi
		elif [[ "$technology" == "splitseq" ]]; then
		barcodefile=${whitelistdir}/splitseq_barcode.txt
		if [[ ! -f ${whitelistdir}/splitseq_barcode.txt ]]; then
		@@ -1680,6 +1692,12 @@ else
		> ${whitelistdir}/sciseq3_barcode.txt
		## to filter unique lines: awk '!a[$0]++' > ${whitelistdir}/sciseq3_barcode.txt
		fi
		elif [[ "$technology" == "scifiseq" ]]; then
		if [[ ! -f ${whitelistdir}/scifi-seq_barcode.txt ]]; then
		#generates all combinations of I1-I2-R1 barcodes
		join -j 9999 ${whitelistdir}/10x_atac_barcodes.txt ${whitelistdir}/ scifi-seq_rt_barcode.txt \| sed "s/ //g" \| \
		> ${whitelistdir}/scifi-seq_barcode.txt
		fi
		elif [[ "$technology" == "splitseq" ]]; then
		#generates all combinations of I1-I2-R1 barcodes
		if [[ ! -f ${whitelistdir}/splitseq_barcode.txt ]]; then
		@@ -2557,15 +2575,16 @@ else
		for convFile in "${convFiles[@]}"; do
		#remove adapter if detected (and keep hairpin/tn5 barcode)
		## TruSeq adapter: ACGACGCTCTTCCGATCT
		cat $convFile \| sed -E '
		sed -E '
		/^ACGACGCTCTTCCGATCT/ {
		s/^ACGACGCTCTTCCGATCT//g
		n
		n
		s/^.{18}//g
		}' \|
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		#remove linker (10 bp barcodes)
		sed -E '
		cat $convFile \| sed -E '
		/^(.{9})CAGAGC/ {
		s/^(.{9})CAGAGC(.{18})/T\1CAGAGC\2/g
		n
		@@ -2602,6 +2621,46 @@ else
		done
		fi

		if [[ "$technology" == "scifiseq" ]]; then
		echo " ...remove adapter for ${technology}"
		for convFile in "${convFiles[@]}"; do
		#remove adapter if detected (and keep hairpin/tn5 barcode)
		## TruSeq adapter: ACGACGCTCTTCCGATCT
		sed -E '
		/^ACACTCTTTCCCTACACGACGCTCTTCCGATCT/ {
		s/^ACACTCTTTCCCTACACGACGCTCTTCCGATCT//g
		n
		n
		s/^.{33}//g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile
		#remove linker and swap RT barcode and UMI (removes one base on either end of barcode): 11 bp barcode not 13 bp
		echo " ...barcode and UMI swapped for ${technology}"
		sed -E '
		/^(.{8}).(.{11)./ {
		s/^(.{8}).(.{11)./\2\1/g
		n
		n
		s/^(.{8}).(.{11)./\2\1/g
		}' $convFile > ${crIN}/.temp
		mv ${crIN}/.temp $convFile

		read=$convFile
		convR1=$read
		convR2=$(echo $read \| perl -pne 's/(.*)_R1/$1_R2/' )
		convI1=$(echo $read \| perl -pne 's/(.*)_R1/$1_I1/' )
		convI2=$(echo $read \| perl -pne 's/(.*)_R1/$1_I2/' )

		echo " ...concatencate 10x ATAC barcodes to R1 from I2 index files"
		# concatenate barcocdes from dual indexes to R1 as (bases 1-16 of the 27 bp) barcode, moving RT barcode (17-27) UMI to (28-35)
		# filter UMI reads by matching tag sequence ATTGCGCAATG (bases 1-11 of R1) and remove as an adapters
		perl sub/ConcatenateDualIndexBarcodes.pl --additive=${convI2} --ref_fastq=${convR1} --out_dir $crIN

		#returns a combined R1 file with I1-I2-R1 concatenated (I1 and I2 are R1 barcode)
		mv $crIN/Concatenated_File.fastq ${convR1}
		done
		fi

		#STRT-Seq
		if [[ "$technology" == "strt-seq" ]] \|\| [[ "$technology" == "strt-seq-c1" ]] \|\| [[ "$technology" == "strt-seq-2i" ]]; then
		echo " ...processsing for ${technology}"

man/launch_universc.sh

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Original line number	Diff line number	Diff line
		@@ -216,6 +216,7 @@ Provides a conversion script to run multiple technologies and custom libraries w
		STRT-Seq-C1 (8 bp barode, 5 bp UMI)
		STRT-Seq-2i (13 bp barcode, 6 bp UMI)
		SmartSeq2 (16 bp barcode, no UMI)
		SCIFI-Seq (27 bp barcode, 8 bp UMI

		A barcode whitelist is provided for all beads or wells for the following technologies:

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