Here we outline the steps involved in creating the pre-built Cell Ranger ARC reference packages. These reference packages can be accessed on the downloads page.
To build your own custom reference for Cell Ranger ARC, see the instructions on this page.
Frankish A, Carbonell-Sala S, Diekhans M, Jungreis I, Loveland JE, Mudge JM, Sisu C, Wright JC, Arnan C, Barnes I, Banerjee A, Bennett R, Berry A, Bignell A, Boix C, Calvet F, Cerdán-Vélez D, Cunningham F, Davidson C, Donaldson S, Dursun C, Fatima R, Giorgetti S, Giron CG, Gonzalez JM, Hardy M, Harrison PW, Hourlier T, Hollis Z, Hunt T, James B, Jiang Y, Johnson R, Kay M, Lagarde J, Martin FJ, Gómez LM, Nair S, Ni P, Pozo F, Ramalingam V, Ruffier M, Schmitt BM, Schreiber JM, Steed E, Suner MM, Sumathipala D, Sycheva I, Uszczynska-Ratajczak B, Wass E, Yang YT, Yates A, Zafrulla Z, Choudhary JS, Gerstein M, Guigo R, Hubbard TJP, Kellis M, Kundaje A, Paten B, Tress ML, Flicek P. Nucleic Acids Res 2023: 51; d1; D942-D949. DOI: 10.1093/nar/gkac1071
Frankish A, Diekhans M, Ferreira AM, Johnson R, Jungreis I, Loveland J, Mudge JM, Sisu C, Wright J, Armstrong J, Barnes I, Berry A, Bignell A, Carbonell Sala S, Chrast J, Cunningham F, Di Domenico T, Donaldson S, Fiddes IT, García Girón C, Gonzalez JM, Grego T, Hardy M, Hourlier T, Hunt T, Izuogu OG, Lagarde J, Martin FJ, Martínez L, Mohanan S, Muir P, Navarro FCP, Parker A, Pei B, Pozo F, Ruffier M, Schmitt BM, Stapleton E, Suner MM, Sycheva I, Uszczynska-Ratajczak B, Xu J, Yates A, Zerbino D, Zhang Y, Aken B, Choudhary JS, Gerstein M, Guigó R, Hubbard TJP, Kellis M, Paten B, Reymond A, Tress ML, Flicek P. Nucleic Acids Res 2019: 47; d1; D766-D773. DOI: 10.1093/nar/gky955
# Genome metadata
genome="GRCh38"
version="2024-A"
# Set up source and build directories
build="${genome}-${version}-build"
mkdir -p "$build"
# Download source files if they do not exist in reference-sources/ folder
source="${genome}-${version}-reference-sources"
mkdir -p "$source"
# Using release 109 for GRCh38 instead of release 110 -- release 110 moved from GRCh38.p13 to GRCh38.p14,
# which unmasked the pseudo-autosomal region. This causes ambiguous mappings to PAR locus genes.
# No other sequence changes were made to the primary assembly.
fasta_url="http://ftp.ensembl.org/pub/release-109/fasta/homo_sapiens/dna/Homo_sapiens.GRCh38.dna.primary_assembly.fa.gz"
fasta_in="${source}/Homo_sapiens.GRCh38.dna.primary_assembly.fa"
gtf_url="http://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_44/gencode.v44.primary_assembly.annotation.gtf.gz"
gtf_in="${source}/gencode.v44.primary_assembly.annotation.gtf"
motifs_url="https://jaspar.elixir.no/download/data/2024/CORE/JASPAR2024_CORE_vertebrates_non-redundant_pfms_jaspar.txt"
motifs_in="${source}/JASPAR2024_CORE_non-redundant_pfms_jaspar.txt"
if [ ! -f "$fasta_in" ]; then
curl -sS "$fasta_url" | zcat > "$fasta_in"
fi
if [ ! -f "$gtf_in" ]; then
curl -sS "$gtf_url" | zcat > "$gtf_in"
fi
if [ ! -f "$motifs_in" ]; then
curl -sS "$motifs_url" > "$motifs_in"
fi
# Modify sequence headers in the Ensembl FASTA to match the file
# "GRCh38.primary_assembly.genome.fa" from GENCODE. Unplaced and unlocalized
# sequences such as "KI270728.1" have the same names in both versions.
#
# Input FASTA:
# >1 dna:chromosome chromosome:GRCh38:1:1:248956422:1 REF
#
# Output FASTA:
# >chr1 1
fasta_modified="$build/$(basename "$fasta_in").modified"
# sed commands:
# 1. Replace metadata after space with original contig name, as in GENCODE
# 2. Add "chr" to names of autosomes and sex chromosomes
# 3. Handle the mitochrondrial chromosome
cat "$fasta_in" \
| sed -E 's/^>(\S+).*/>\1 \1/' \
| sed -E 's/^>([0-9]+|[XY]) />chr\1 /' \
| sed -E 's/^>MT />chrM /' \
> "$fasta_modified"
# Remove version suffix from transcript, gene, and exon IDs in order to match
# previous Cell Ranger reference packages
#
# Input GTF:
# ... gene_id "ENSG00000223972.5"; ...
# Output GTF:
# ... gene_id "ENSG00000223972"; gene_version "5"; ...
gtf_modified="$build/$(basename "$gtf_in").modified"
# Pattern matches Ensembl gene, transcript, and exon IDs for human or mouse:
ID="(ENS(MUS)?[GTE][0-9]+)\.([0-9]+)"
cat "$gtf_in" \
| sed -E 's/gene_id "'"$ID"'";/gene_id "\1"; gene_version "\3";/' \
| sed -E 's/transcript_id "'"$ID"'";/transcript_id "\1"; transcript_version "\3";/' \
| sed -E 's/exon_id "'"$ID"'";/exon_id "\1"; exon_version "\3";/' \
> "$gtf_modified"
# Define string patterns for GTF tags
# NOTES:
# Since Ensembl 110, polymorphic pseudogenes are now just protein_coding.
# Readthrough genes are annotated with the readthrough_transcript tag.
BIOTYPE_PATTERN=\
"(protein_coding|protein_coding_LoF|lncRNA|\
IG_C_gene|IG_D_gene|IG_J_gene|IG_LV_gene|IG_V_gene|\
IG_V_pseudogene|IG_J_pseudogene|IG_C_pseudogene|\
TR_C_gene|TR_D_gene|TR_J_gene|TR_V_gene|\
TR_V_pseudogene|TR_J_pseudogene)"
GENE_PATTERN="gene_type \"${BIOTYPE_PATTERN}\""
TX_PATTERN="transcript_type \"${BIOTYPE_PATTERN}\""
READTHROUGH_PATTERN="tag \"readthrough_transcript\""
# Construct the gene ID allowlist. We filter the list of all transcripts
# based on these criteria:
# - allowable gene_type (biotype)
# - allowable transcript_type (biotype)
# - no "readthrough_transcript" tag
# We then collect the list of gene IDs that have at least one associated
# transcript passing the filters.
cat "$gtf_modified" \
| awk '$3 == "transcript"' \
| grep -E "$GENE_PATTERN" \
| grep -E "$TX_PATTERN" \
| grep -Ev "$READTHROUGH_PATTERN" \
| sed -E 's/.*(gene_id "[^"]+").*/\1/' \
| sort \
| uniq \
> "${build}/gene_allowlist"
# NOTES:
# Since Ensembl 110, the PAR locus genes are included on chrY as copies of chrX
# Using the GRCh38.p13 assembly hard masks these regions on chrY, but removing the
# chrY PAR genes is still desirable so they do not end up as extra entries in the output.
# The awk command below excludes all PAR_Y genes, including XGY2.
# The non-coding gene XGY2 straddles the PAR1 boundary on chrY, and is homologous to XG on chrX.
# GRCh38-2024-A excludes XGY2, but includes SRY and ENSG00000286130, which are in an intron of XGY2,
# and RPS4Y1, which overlaps XGY2.
# Filter the GTF file based on the gene allowlist
gtf_filtered="${build}/$(basename "$gtf_in").filtered"
# Copy header lines beginning with "#"
grep -E "^#" "$gtf_modified" > "$gtf_filtered"
# Filter to the gene allowlist, and then remove PAR_Y genes
grep -Ff "${build}/gene_allowlist" "$gtf_modified" \
| awk -F "\t" '$1 != "chrY" || $1 == "chrY" && $4 >= 2752083 && $4 < 56887903 && !/ENSG00000290840/' \
>> "$gtf_filtered"
# Change motif headers so the human-readable motif name precedes the motif
# identifier. So ">MA0004.1 Arnt" -> ">Arnt_MA0004.1".
motifs_modified="$build/$(basename "$motifs_in").modified"
awk '{
if ( substr($1, 1, 1) == ">" ) {
print ">" $2 "_" substr($1,2)
} else {
print
}
}' "$motifs_in" > "$motifs_modified"
# Create a config file
config_in="${build}/config"
echo """{
organism: \"Homo_sapiens\"
genome: [\""$genome"\"]
input_fasta: [\""$fasta_modified"\"]
input_gtf: [\""$gtf_filtered"\"]
input_motifs: \""$motifs_modified"\"
non_nuclear_contigs: [\"chrM\"]
}""" > "$config_in"
# Create reference package
cellranger-arc mkref --ref-version="$version" \
--config="$config_in" --nthreads=16
# Genome metadata
genome="GRCm39"
version="2024-A"
# Set up source and build directories
build="${genome}-${version}-build"
mkdir -p "$build"
# Download source files if they do not exist in reference-sources/ folder
source="${genome}-${version}-reference-sources"
mkdir -p "$source"
fasta_url="http://ftp.ensembl.org/pub/release-110/fasta/mus_musculus/dna/Mus_musculus.GRCm39.dna.primary_assembly.fa.gz"
fasta_in="${source}/Mus_musculus.GRCm39.dna.primary_assembly.fa"
gtf_url="http://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_mouse/release_M33/gencode.vM33.primary_assembly.annotation.gtf.gz"
gtf_in="${source}/gencode.vM33.primary_assembly.annotation.gtf"
motifs_url="https://jaspar.elixir.no/download/data/2024/CORE/JASPAR2024_CORE_vertebrates_non-redundant_pfms_jaspar.txt"
motifs_in="${source}/JASPAR2024_CORE_non-redundant_pfms_jaspar.txt"
if [ ! -f "$fasta_in" ]; then
curl -sS "$fasta_url" | zcat > "$fasta_in"
fi
if [ ! -f "$gtf_in" ]; then
curl -sS "$gtf_url" | zcat > "$gtf_in"
fi
if [ ! -f "$motifs_in" ]; then
curl -sS "$motifs_url" > "$motifs_in"
fi
# Modify sequence headers in the Ensembl FASTA to match the file
# "GRCm38.primary_assembly.genome.fa" from GENCODE. Unplaced and unlocalized
# sequences such as "GL456210.1" have the same names in both versions.
#
# Input FASTA:
# >1 dna:chromosome chromosome:GRCm38:1:1:195471971:1 REF
#
# Output FASTA:
# >chr1 1
fasta_modified="$build/$(basename "$fasta_in").modified"
# sed commands:
# 1. Replace metadata after space with original contig name, as in GENCODE
# 2. Add "chr" to names of autosomes and sex chromosomes
# 3. Handle the mitochrondrial chromosome
cat "$fasta_in" \
| sed -E 's/^>(\S+).*/>\1 \1/' \
| sed -E 's/^>([0-9]+|[XY]) />chr\1 /' \
| sed -E 's/^>MT />chrM /' \
> "$fasta_modified"
# Remove version suffix from transcript, gene, and exon IDs in order to match
# previous Cell Ranger reference packages
#
# Input GTF:
# ... gene_id "ENSMUSG00000102693.1"; ...
# Output GTF:
# ... gene_id "ENSMUSG00000102693"; gene_version "1"; ...
gtf_modified="$build/$(basename "$gtf_in").modified"
# Pattern matches Ensembl gene, transcript, and exon IDs for human or mouse:
ID="(ENS(MUS)?[GTE][0-9]+)\.([0-9]+)"
cat "$gtf_in" \
| sed -E 's/gene_id "'"$ID"'";/gene_id "\1"; gene_version "\3";/' \
| sed -E 's/transcript_id "'"$ID"'";/transcript_id "\1"; transcript_version "\3";/' \
| sed -E 's/exon_id "'"$ID"'";/exon_id "\1"; exon_version "\3";/' \
> "$gtf_modified"
# Define string patterns for GTF tags
# Since Ensembl 110, polymorphic pseudogenes are now just protein_coding.
# Readthrough genes are annotated with the readthrough_transcript tag.
BIOTYPE_PATTERN=\
"(protein_coding|protein_coding_LoF|lncRNA|\
IG_C_gene|IG_D_gene|IG_J_gene|IG_LV_gene|IG_V_gene|\
IG_V_pseudogene|IG_J_pseudogene|IG_C_pseudogene|\
TR_C_gene|TR_D_gene|TR_J_gene|TR_V_gene|\
TR_V_pseudogene|TR_J_pseudogene)"
GENE_PATTERN="gene_type \"${BIOTYPE_PATTERN}\""
TX_PATTERN="transcript_type \"${BIOTYPE_PATTERN}\""
READTHROUGH_PATTERN="tag \"readthrough_transcript\""
# Construct the gene ID allowlist. We filter the list of all transcripts
# based on these criteria:
# - allowable gene_type (biotype)
# - allowable transcript_type (biotype)
# - no "readthrough_transcript" tag
# We then collect the list of gene IDs that have at least one associated
# transcript passing the filters.
cat "$gtf_modified" \
| awk '$3 == "transcript"' \
| grep -E "$GENE_PATTERN" \
| grep -E "$TX_PATTERN" \
| grep -Ev "$READTHROUGH_PATTERN" \
| sed -E 's/.*(gene_id "[^"]+").*/\1/' \
| sort \
| uniq \
> "${build}/gene_allowlist"
# Filter the GTF file based on the gene allowlist
gtf_filtered="${build}/$(basename "$gtf_in").filtered"
# Copy header lines beginning with "#"
grep -E "^#" "$gtf_modified" > "$gtf_filtered"
# Filter to the gene allowlist
grep -Ff "${build}/gene_allowlist" "$gtf_modified" \
>> "$gtf_filtered"
# Change motif headers so the human-readable motif name precedes the motif
# identifier. So ">MA0004.1 Arnt" -> ">Arnt_MA0004.1".
motifs_modified="$build/$(basename "$motifs_in").modified"
awk '{
if ( substr($1, 1, 1) == ">" ) {
print ">" $2 "_" substr($1,2)
} else {
print
}
}' "$motifs_in" > "$motifs_modified"
# Create a config file
config_in="${build}/config"
echo """{
organism: \"Mus_musculus\"
genome: [\""$genome"\"]
input_fasta: [\""$fasta_modified"\"]
input_gtf: [\""$gtf_filtered"\"]
input_motifs: \""$motifs_modified"\"
non_nuclear_contigs: [\"chrM\"]
}""" > "$config_in"
# Create reference package
cellranger-arc mkref --ref-version="$version" \
--config="$config_in" --nthreads=16
# Genome metadata
genome="GRCh38"
version="2020-A"
# Set up source and build directories
build="${genome}-${version}-build"
mkdir -p "$build"
# Download source files if they do not exist in reference-sources/ folder
source="${genome}-${version}-reference-sources"
mkdir -p "$source"
fasta_url="http://ftp.ensembl.org/pub/release-98/fasta/homo_sapiens/dna/Homo_sapiens.GRCh38.dna.primary_assembly.fa.gz"
fasta_in="${source}/Homo_sapiens.GRCh38.dna.primary_assembly.fa"
gtf_url="http://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_32/gencode.v32.primary_assembly.annotation.gtf.gz"
gtf_in="${source}/gencode.v32.primary_assembly.annotation.gtf"
motifs_url="https://jaspar.genereg.net/download/data/2018/CORE/JASPAR2018_CORE_vertebrates_non-redundant_pfms_jaspar.txt"
motifs_in="${source}/JASPAR2018_CORE_non-redundant_pfms_jaspar.txt"
if [ ! -f "$fasta_in" ]; then
curl -sS "$fasta_url" | zcat > "$fasta_in"
fi
if [ ! -f "$gtf_in" ]; then
curl -sS "$gtf_url" | zcat > "$gtf_in"
fi
if [ ! -f "$motifs_in" ]; then
curl -sS "$motifs_url" > "$motifs_in"
fi
# Modify sequence headers in the Ensembl FASTA to match the file
# "GRCh38.primary_assembly.genome.fa" from GENCODE. Unplaced and unlocalized
# sequences such as "KI270728.1" have the same names in both versions.
#
# Input FASTA:
# >1 dna:chromosome chromosome:GRCh38:1:1:248956422:1 REF
#
# Output FASTA:
# >chr1 1
fasta_modified="$build/$(basename "$fasta_in").modified"
# sed commands:
# 1. Replace metadata after space with original contig name, as in GENCODE
# 2. Add "chr" to names of autosomes and sex chromosomes
# 3. Handle the mitochrondrial chromosome
cat "$fasta_in" \
| sed -E 's/^>(\S+).*/>\1 \1/' \
| sed -E 's/^>([0-9]+|[XY]) />chr\1 /' \
| sed -E 's/^>MT />chrM /' \
> "$fasta_modified"
# Remove version suffix from transcript, gene, and exon IDs in order to match
# previous Cell Ranger reference packages
#
# Input GTF:
# ... gene_id "ENSG00000223972.5"; ...
# Output GTF:
# ... gene_id "ENSG00000223972"; gene_version "5"; ...
gtf_modified="$build/$(basename "$gtf_in").modified"
# Pattern matches Ensembl gene, transcript, and exon IDs for human or mouse:
ID="(ENS(MUS)?[GTE][0-9]+)\.([0-9]+)"
cat "$gtf_in" \
| sed -E 's/gene_id "'"$ID"'";/gene_id "\1"; gene_version "\3";/' \
| sed -E 's/transcript_id "'"$ID"'";/transcript_id "\1"; transcript_version "\3";/' \
| sed -E 's/exon_id "'"$ID"'";/exon_id "\1"; exon_version "\3";/' \
> "$gtf_modified"
# Define string patterns for GTF tags
# NOTES:
# - Since GENCODE release 31/M22 (Ensembl 97), the "lncRNA" and "antisense"
# biotypes are part of a more generic "lncRNA" biotype.
# - These filters are relevant only to GTF files from GENCODE. The GTFs from
# Ensembl release 98 have the following differences:
# - The names "gene_biotype" and "transcript_biotype" are used instead of
# "gene_type" and "transcript_type".
# - Readthrough transcripts are present but are not marked with the
# "readthrough_transcript" tag.
# - Only the X chromosome versions of genes in the pseudoautosomal regions
# are present, so there is no "PAR" tag.
BIOTYPE_PATTERN=\
"(protein_coding|lncRNA|\
IG_C_gene|IG_D_gene|IG_J_gene|IG_LV_gene|IG_V_gene|\
IG_V_pseudogene|IG_J_pseudogene|IG_C_pseudogene|\
TR_C_gene|TR_D_gene|TR_J_gene|TR_V_gene|\
TR_V_pseudogene|TR_J_pseudogene)"
GENE_PATTERN="gene_type \"${BIOTYPE_PATTERN}\""
TX_PATTERN="transcript_type \"${BIOTYPE_PATTERN}\""
READTHROUGH_PATTERN="tag \"readthrough_transcript\""
PAR_PATTERN="tag \"PAR\""
# Construct the gene ID allowlist. We filter the list of all transcripts
# based on these criteria:
# - allowable gene_type (biotype)
# - allowable transcript_type (biotype)
# - no "PAR" tag (only present for Y chromosome PAR)
# - no "readthrough_transcript" tag
# We then collect the list of gene IDs that have at least one associated
# transcript passing the filters.
cat "$gtf_modified" \
| awk '$3 == "transcript"' \
| grep -E "$GENE_PATTERN" \
| grep -E "$TX_PATTERN" \
| grep -Ev "$READTHROUGH_PATTERN" \
| grep -Ev "$PAR_PATTERN" \
| sed -E 's/.*(gene_id "[^"]+").*/\1/' \
| sort \
| uniq \
> "${build}/gene_allowlist"
# Filter the GTF file based on the gene allowlist
gtf_filtered="${build}/$(basename "$gtf_in").filtered"
# Copy header lines beginning with "#"
grep -E "^#" "$gtf_modified" > "$gtf_filtered"
# Filter to the gene allowlist
grep -Ff "${build}/gene_allowlist" "$gtf_modified" \
>> "$gtf_filtered"
# Change motif headers so the human-readable motif name precedes the motif
# identifier. So ">MA0004.1 Arnt" -> ">Arnt_MA0004.1".
motifs_modified="$build/$(basename "$motifs_in").modified"
awk '{
if ( substr($1, 1, 1) == ">" ) {
print ">" $2 "_" substr($1,2)
} else {
print
}
}' "$motifs_in" > "$motifs_modified"
# Create a config file
config_in="${build}/config"
echo """{
organism: \"Homo_sapiens\"
genome: [\""$genome"\"]
input_fasta: [\""$fasta_modified"\"]
input_gtf: [\""$gtf_filtered"\"]
input_motifs: \""$motifs_modified"\"
non_nuclear_contigs: [\"chrM\"]
}""" > "$config_in"
# Create reference package
cellranger-arc mkref --ref-version="$version" \
--config="$config_in"
# Genome metadata
genome="mm10"
version="2020-A"
# Set up source and build directories
build="${genome}-${version}-build"
mkdir -p "$build"
# Download source files if they do not exist in reference_sources/ folder
source="${genome}-${version}-reference-sources"
mkdir -p "$source"
fasta_url="http://ftp.ensembl.org/pub/release-98/fasta/mus_musculus/dna/Mus_musculus.GRCm38.dna.primary_assembly.fa.gz"
fasta_in="${source}/Mus_musculus.GRCm38.dna.primary_assembly.fa"
gtf_url="http://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_mouse/release_M23/gencode.vM23.primary_assembly.annotation.gtf.gz"
gtf_in="${source}/gencode.vM23.primary_assembly.annotation.gtf"
motifs_url="https://jaspar.genereg.net/download/data/2018/CORE/JASPAR2018_CORE_vertebrates_non-redundant_pfms_jaspar.txt"
motifs_in="${source}/JASPAR2018_CORE_vertebrates_non-redundant_pfms_jaspar.txt"
if [ ! -f "$fasta_in" ]; then
curl -sS "$fasta_url" | zcat > "$fasta_in"
fi
if [ ! -f "$gtf_in" ]; then
curl -sS "$gtf_url" | zcat > "$gtf_in"
fi
if [ ! -f "$motifs_in" ]; then
curl -sS "$motifs_url" > "$motifs_in"
fi
# Modify sequence headers in the Ensembl FASTA to match the file
# "GRCm38.primary_assembly.genome.fa" from GENCODE. Unplaced and unlocalized
# sequences such as "GL456210.1" have the same names in both versions.
#
# Input FASTA:
# >1 dna:chromosome chromosome:GRCm38:1:1:195471971:1 REF
#
# Output FASTA:
# >chr1 1
fasta_modified="$build/$(basename "$fasta_in").modified"
# sed commands:
# 1. Replace metadata after space with original contig name, as in GENCODE
# 2. Add "chr" to names of autosomes and sex chromosomes
# 3. Handle the mitochrondrial chromosome
cat "$fasta_in" \
| sed -E 's/^>(\S+).*/>\1 \1/' \
| sed -E 's/^>([0-9]+|[XY]) />chr\1 /' \
| sed -E 's/^>MT />chrM /' \
> "$fasta_modified"
# Remove version suffix from transcript, gene, and exon IDs in order to match
# previous Cell Ranger reference packages
#
# Input GTF:
# ... gene_id "ENSMUSG00000102693.1"; ...
# Output GTF:
# ... gene_id "ENSMUSG00000102693"; gene_version "1"; ...
gtf_modified="$build/$(basename "$gtf_in").modified"
# Pattern matches Ensembl gene, transcript, and exon IDs for human or mouse:
ID="(ENS(MUS)?[GTE][0-9]+)\.([0-9]+)"
cat "$gtf_in" \
| sed -E 's/gene_id "'"$ID"'";/gene_id "\1"; gene_version "\3";/' \
| sed -E 's/transcript_id "'"$ID"'";/transcript_id "\1"; transcript_version "\3";/' \
| sed -E 's/exon_id "'"$ID"'";/exon_id "\1"; exon_version "\3";/' \
> "$gtf_modified"
# Define string patterns for GTF tags
# NOTES:
# - Since GENCODE release 31/M22 (Ensembl 97), the "lncRNA" and "antisense"
# biotypes are part of a more generic "lncRNA" biotype.
# - These filters are relevant only to GTF files from GENCODE. The GTFs from
# Ensembl release 98 have the following differences:
# - The names "gene_biotype" and "transcript_biotype" are used instead of
# "gene_type" and "transcript_type".
# - Readthrough transcripts are present but are not marked with the
# "readthrough_transcript" tag.
BIOTYPE_PATTERN=\
"(protein_coding|lncRNA|\
IG_C_gene|IG_D_gene|IG_J_gene|IG_LV_gene|IG_V_gene|\
IG_V_pseudogene|IG_J_pseudogene|IG_C_pseudogene|\
TR_C_gene|TR_D_gene|TR_J_gene|TR_V_gene|\
TR_V_pseudogene|TR_J_pseudogene)"
GENE_PATTERN="gene_type \"${BIOTYPE_PATTERN}\""
TX_PATTERN="transcript_type \"${BIOTYPE_PATTERN}\""
READTHROUGH_PATTERN="tag \"readthrough_transcript\""
# Construct the gene ID allowlist. We filter the list of all transcripts
# based on these criteria:
# - allowable gene_type (biotype)
# - allowable transcript_type (biotype)
# - no "readthrough_transcript" tag
# We then collect the list of gene IDs that have at least one associated
# transcript passing the filters.
cat "$gtf_modified" \
| awk '$3 == "transcript"' \
| grep -E "$GENE_PATTERN" \
| grep -E "$TX_PATTERN" \
| grep -Ev "$READTHROUGH_PATTERN" \
| sed -E 's/.*(gene_id "[^"]+").*/\1/' \
| sort \
| uniq \
> "${build}/gene_allowlist"
# Filter the GTF file based on the gene allowlist
gtf_filtered="${build}/$(basename "$gtf_in").filtered"
# Copy header lines beginning with "#"
grep -E "^#" "$gtf_modified" > "$gtf_filtered"
# Filter to the gene allowlist
grep -Ff "${build}/gene_allowlist" "$gtf_modified" \
>> "$gtf_filtered"
# Change motif headers so the human-readable motif name precedes the motif
# identifier. So ">MA0004.1 Arnt" -> ">Arnt_MA0004.1".
motifs_modified="$build/$(basename "$motifs_in").modified"
awk '{
if ( substr($1, 1, 1) == ">" ) {
print ">" $2 "_" substr($1,2)
} else {
print
}
}' "$motifs_in" > "$motifs_modified"
# Create a config file
config_in="${build}/config"
echo """{
organism: \"Mus_musculus\"
genome: [\""$genome"\"]
input_fasta: [\""$fasta_modified"\"]
input_gtf: [\""$gtf_filtered"\"]
input_motifs: \""$motifs_modified"\"
non_nuclear_contigs: [\"chrM\"]
}""" > "$config_in"
# Create reference package
cellranger-arc mkref --ref-version="$version" \
--config="$config_in"