diff --git a/README.md b/README.md
index deeeda1e13ce7bfd0918b9bef35dbfd72b4fbb0e..6bd20fabeba8a3b4b0075e7df972a48c58db9b8b 100644
--- a/README.md
+++ b/README.md
@@ -168,6 +168,9 @@ files should look like, specifically:
- [samples.tsv](tests/input_files/samples.tsv)
- [config.yaml](tests/input_files/config.yaml)
+ - For more details and explanations, refer to the [pipeline-documentation]
+
+
4. Create a runner script. Pick one of the following choices for either local
or cluster execution. Before execution of the respective command, you need to
remember to update the argument of the `--singularity-args` option of a
diff --git a/pipeline_documentation.md b/pipeline_documentation.md
index bbef50f59eeb32d5da838b9d80b0bc91c6e18d49..070b3662f495968f869f5e7618317612d7f95b6e 100644
--- a/pipeline_documentation.md
+++ b/pipeline_documentation.md
@@ -119,8 +119,7 @@ mean | Required for [kallisto](#third-party-software-used) and [Salmon](#third-p
multimappers | Required for [STAR](#third-party-software-used). Maximum number of multiple alignments allowed for a read; if exceeded, the read is considered unmapped. | `int`
soft_clip | Required for [STAR](#third-party-software-used). One of `Local` (standard local alignment with soft-clipping allowed) or `EndToEnd` (force end-to-end read alignment, do not soft-clip). | `str`
pass_mode | Required for [STAR](#third-party-software-used). One of `None` (1-pass mapping) or `Basic` (basic 2-pass mapping, with all 1st-pass junctions inserted into the genome indices on the fly). | `str`
-libtype | Required for [Salmon](#third-party-software-used). See [Salmon manual][docs-salmon] for allowed values. If in doubt, enter `A` to automatically infer the library type. | `str`
-kallisto_directionality | Required for [kallisto](#third-party-software-used) and [ALFA](#third-party-software-used). One of `--fr-stranded` (strand-specific reads, first read forward) and `--rf-stranded` (strand-specific reads, first read reverse) | `str`
+libtype | Required for [Salmon](#third-party-software-used), and, after internal conversion, for [kallisto](#third-party-software-used) and [ALFA](#third-party-software-used) . See [Salmon manual][docs-salmon] for allowed values. **WARNING**: do *NOT* use `A` to automatically infer the salmon library type, this will cause kallisto and ALFA to fail. | `str`
fq1_polya3p | Required for [Cutadapt](#third-party-software-used). Stretch of `A`s or `T`s, depending on read orientation. Trimmed from the 3' end of the read. Use value such as `XXXXXXXXXXXXXXX` if no poly(A) stretch present or if no trimming is desired. | `str`
fq1_polya5p | Required for [Cutadapt](#third-party-software-used). Stretch of `A`s or `T`s, depending on read orientation. Trimmed from the 5' end of the read. Use value such as `XXXXXXXXXXXXXXX` if no poly(A) stretch present or if no trimming is desired. | `str`
fq2_polya3p | Required for [Cutadapt](#third-party-software-used). Stretch of `A`s or `T`s, depending on read orientation. Trimmed from the 3' end of the read. Use value such as `XXXXXXXXXXXXXXX` if no poly(A) stretch present or if no trimming is desired. Value ignored for single-end libraries. | `str`
@@ -299,7 +298,7 @@ Rename and copy stranded bedGraph coverage tracks such that they comply with
> Local rule
>
> Renaming to `plus.bg` and `minus.bg` depends on library orientation, which is
-> provided by user in sample table column `kallisto_directionality`.
+> provided by user in sample table column `libtype`.
- **Input**
- Coverage file (`.bg`); from [**star_rpm**](#star_rpm)
@@ -465,15 +464,15 @@ Create index for [**ALFA**](#third-party-software-used).
Annotate alignments with [**ALFA**](#third-party-software-used).
-> For details on output plots, see [ALFA documentation][docs-alfa].
+> For details on output plots, see [ALFA documentation][docs-alfa].
+> Note: the read orientation of a sample will be inferred from salmon `libtype` specified in `samples.tsv`
- **Input**
- Coverage files, renamed (`.bg`); from
[**rename_star_rpm_for_alfa**](#rename_star_rpm_for_alfa)
- ALFA index, stranded; from [**generate_alfa_index**](#generate_alfa_index)
- **Parameters**
- - **samples.tsv**
- - `-s`: library orientation; specified by user in sample table column `kallisto_directionality`
+
- **Output**
- Figures for biotypes and feature categories (`.pdf`)
- Feature counts table (custom `.tsv`); used in
@@ -665,6 +664,7 @@ Estimate transcript- and gene-level expression with
Generate pseudoalignments of reads to transcripts with
[**kallisto**](#third-party-software-used).
+> Note: the kallisto strandedness parameter will be inferred from salmon `libtype` specified in `samples.tsv`
- **Input**
- Reads file (`.fastq.gz`); from
@@ -672,7 +672,6 @@ Generate pseudoalignments of reads to transcripts with
- Index; from [**create_index_kallisto**](#create_index_kallisto)
- **Parameters**
- **samples.tsv**
- - `directionality`; specify in sample table column `kallisto_directionality`
- `-l`: mean of distribution of fragment lengths; specify in sample table column `mean` **(single-end only)**
- `-s`: standard deviation of distribution of fragment lengths; specify in sample table column `sd` **(single-end only)**
- **Output**
diff --git a/scripts/prepare_inputs.py b/scripts/prepare_inputs.py
index 426902a014801bd3ddc677d9b6f01d644f8e8334..bc164aa1a296cde32816a65cf614417ed9c8fa02 100755
--- a/scripts/prepare_inputs.py
+++ b/scripts/prepare_inputs.py
@@ -137,9 +137,11 @@ def parse_cli_args() -> argparse.Namespace:
behavior.add_argument(
"--libtype",
type=str,
- default="A",
- help="Salmon: library type (default: %(default)s)",
+ default="",
+ help="Salmon library type (default: %(default)s). Leave empty to infer one of 'SF', 'SR', 'ISF', 'ISR'."
+ "Warning: If value is provided by user, it will be applied to ALL samples. If the table contains samples from different sequencing modes this might cause errors in zarp.",
metavar="STR",
+ choices=["", "SF", "SR", "ISF", "ISR", "OSF", "OSR", "MSF", "MSR"]
)
report = parser.add_argument_group("report")
@@ -311,72 +313,37 @@ def kmer_from_read_length(
return k
-def get_strand_param_kallisto(directionality: str) -> str:
+def get_libtype(directionality: str, seqmode: str) -> str:
"""
- Returns appropriate strand info parameter for kallisto
- (https://pachterlab.github.io/kallisto/), given a string indicating the
- "directionality" of a sequencing library.
+ Returns libtype (https://salmon.readthedocs.io/en/latest/library_type.html) given strings indicating the
+ "directionality", and sequencing mode of a sequencing library, respectively.
:param directionality: direction in which library was sequenced (one of
"SENSE" and "ANTISENSE")
- :returns: appropriate kallisto option for specified directionality; an
- empty string is returned if the directionality value is empty or not
- recognized
- """
- if directionality == "SENSE":
- option = "--fr"
- elif directionality == "ANTISENSE":
- option = "--rf"
- else:
- option = ""
- return option
-
+ :param seqmode: sequencing mode(one of
+ "pe" and "se")
-def get_strand_param_alfa(directionality: str) -> str:
+ :returns: salmon code (one of 'SF', 'SR', 'ISF', 'ISR') for specified directionality;
"""
- Returns appropriate strand info parameter for ALFA
- (https://github.com/biocompibens/ALFA), given a string indicating the
- "directionality" of a sequencing library.
- :param directionality: direction in which library was sequenced (one of
- "SENSE" and "ANTISENSE")
-
- :returns: appropriate ALFA option for specified directionality; an empty
- string is returned if the directionality value is empty or not
- recognized
- """
- if directionality == 'SENSE':
- option = 'fr-firststrand'
- elif directionality == 'ANTISENSE':
- option = 'fr-secondstrand'
+ if seqmode == "pe":
+ option = "I"
else:
- option = ''
- return option
+ option = ""
-
-def get_strand_names_alfa(directionality: str) -> Tuple[str, str]:
- """
- Returns appropriate strand name suffixes for ALFA
- (https://github.com/biocompibens/ALFA), given a string indicating the
- "directionality" of a sequencing library.
-
- :param directionality: direction in which library was sequenced (one of
- "SENSE" and "ANTISENSE")
-
- :returns: tuple of ALFA strand name suffixes for two coverage tracks of a
- paired-end sequencing library
- """
if directionality == "SENSE":
- plus = "str1"
- minus = "str2"
+ option += "SF"
elif directionality == "ANTISENSE":
- minus = "str1"
- plus = "str2"
+ option += "SR"
else:
- plus = ""
- minus = ""
- return (plus, minus)
+ logger.error(
+ f"[ERROR] Can't infer library type."
+ f"Make sure directionality and sequencing mode are specified correctly."
+ )
+ sys.exit("Execution aborted.")
+
+ return option
def get_polya_adapter_seqs(directionality: str) -> Tuple[str, str]:
@@ -583,9 +550,6 @@ def main(args):
mean = lk_mean
fq1_polya_3p, fq1_polya_5p = get_polya_adapter_seqs(lk_mate1_direction)
fq2_polya_3p, fq2_polya_5p = get_polya_adapter_seqs(lk_mate2_direction)
- kallisto_directionality = get_strand_param_kallisto(lk_mate1_direction)
- alfa_directionality = get_strand_param_alfa(lk_mate1_direction)
- alfa_plus, alfa_minus = get_strand_names_alfa(lk_mate1_direction)
# construct row in Snakemake input table
snakemake_table.loc[index, 'sample'] = sample
@@ -603,17 +567,20 @@ def main(args):
snakemake_table.loc[index, 'genome'] = genome
snakemake_table.loc[index, 'sd'] = sd
snakemake_table.loc[index, 'mean'] = mean
- snakemake_table.loc[index, 'kallisto_directionality'] = \
- kallisto_directionality
- snakemake_table.loc[index, 'alfa_directionality'] = alfa_directionality
- snakemake_table.loc[index, 'alfa_plus'] = alfa_plus
- snakemake_table.loc[index, 'alfa_minus'] = alfa_minus
# add CLI argument-dependent parameters
snakemake_table.loc[index, 'multimappers'] = args.multimappers
snakemake_table.loc[index, 'soft_clip'] = args.soft_clip
snakemake_table.loc[index, 'pass_mode'] = args.pass_mode
- snakemake_table.loc[index, 'libtype'] = args.libtype
+
+ if not args.libtype:
+ snakemake_table.loc[index, 'libtype'] = get_libtype(lk_mate1_direction, seqmode)
+ elif args.libtype in ['SF', 'SR', 'ISF', 'ISR', 'OSF', 'OSR', 'MSF', 'MSR']:
+ snakemake_table.loc[index, 'libtype'] = args.libtype
+ logger.warning(
+ f"Library type {args.libtype} set for sample {sample}."
+ )
+
if args.trim_polya is True:
snakemake_table.loc[index, 'fq1_polya_3p'] = fq1_polya_3p
snakemake_table.loc[index, 'fq1_polya_5p'] = fq1_polya_5p
diff --git a/tests/input_files/samples.multiple_lanes.tsv b/tests/input_files/samples.multiple_lanes.tsv
index 7968fb46b8331974de83555f0ea6d01b0fe08821..6b32ab8b07181b80013eaff99b3abd1be12f0982 100644
--- a/tests/input_files/samples.multiple_lanes.tsv
+++ b/tests/input_files/samples.multiple_lanes.tsv
@@ -1,5 +1,5 @@
-sample seqmode fq1 fq2 index_size kmer fq1_3p fq1_5p fq2_3p fq2_5p organism gtf genome sd mean kallisto_directionality alfa_directionality alfa_plus alfa_minus multimappers soft_clip pass_mode libtype fq1_polya_3p fq1_polya_5p fq2_polya_3p fq2_polya_5p
-synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/pe_lane1/synthetic_split_lane1.mate_1.fastq.gz ../input_files/pe_lane1/synthetic_split_lane1.mate_2.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 --fr fr-firststrand str1 str2 10 EndToEnd None A AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX TTTTTTTTTTTTTTT
-synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/pe_lane2/synthetic_split_lane2.mate_1.fastq.gz ../input_files/pe_lane2/synthetic_split_lane2.mate_2.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 --fr fr-firststrand str1 str2 10 EndToEnd None A AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX TTTTTTTTTTTTTTT
-synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/se_lane1/synthetic_split_lane1.mate_1.fastq.gz XXXXXXXXXXXXXXX 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 --fr fr-firststrand str1 str2 10 EndToEnd None A AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX
-synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/se_lane2/synthetic_split_lane2.mate_1.fastq.gz XXXXXXXXXXXXXXX 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 --fr fr-firststrand str1 str2 10 EndToEnd None A AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX
+sample seqmode fq1 fq2 index_size kmer fq1_3p fq1_5p fq2_3p fq2_5p organism gtf genome sd mean multimappers soft_clip pass_mode libtype fq1_polya_3p fq1_polya_5p fq2_polya_3p fq2_polya_5p
+synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/pe_lane1/synthetic_split_lane1.mate_1.fastq.gz ../input_files/pe_lane1/synthetic_split_lane1.mate_2.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None ISF AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX TTTTTTTTTTTTTTT
+synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/pe_lane2/synthetic_split_lane2.mate_1.fastq.gz ../input_files/pe_lane2/synthetic_split_lane2.mate_2.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None ISF AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX TTTTTTTTTTTTTTT
+synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/se_lane1/synthetic_split_lane1.mate_1.fastq.gz XXXXXXXXXXXXXXX 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None SF AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX
+synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/se_lane2/synthetic_split_lane2.mate_1.fastq.gz XXXXXXXXXXXXXXX 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None SF AAAAAAAAAAAAAAA XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX XXXXXXXXXXXXXXX
diff --git a/tests/input_files/samples.tsv b/tests/input_files/samples.tsv
index 4b2630287454e8a7d4dbc7652f1897410a905ef5..58ac18fdfd3d0a741bdce639b03c99cc7fd0a313 100644
--- a/tests/input_files/samples.tsv
+++ b/tests/input_files/samples.tsv
@@ -1,3 +1,3 @@
-sample seqmode fq1 index_size kmer fq1_3p fq1_5p organism gtf genome sd mean multimappers soft_clip pass_mode libtype fq1_polya_3p fq1_polya_5p kallisto_directionality alfa_directionality alfa_plus alfa_minus fq2 fq2_3p fq2_5p fq2_polya_3p fq2_polya_5p
-synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/project1/synthetic.mate_1.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None A AAAAAAAAAAAAAAAAA XXXXXXXXXXXXXXXXX --fr fr-firststrand str1 str2 ../input_files/project1/synthetic.mate_2.fastq.gz AGATCGGAAGAGCGT XXXXXXXXXXXXX XXXXXXXXXXXXXXXXX TTTTTTTTTTTTTTTTT
-synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/project2/synthetic.mate_1.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None A AAAAAAAAAAAAAAAAA XXXXXXXXXXXXXXXXX --fr fr-firststrand str1 str2 XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX
+sample seqmode fq1 index_size kmer fq1_3p fq1_5p organism gtf genome sd mean multimappers soft_clip pass_mode libtype fq1_polya_3p fq1_polya_5p fq2 fq2_3p fq2_5p fq2_polya_3p fq2_polya_5p
+synthetic_10_reads_paired_synthetic_10_reads_paired pe ../input_files/project1/synthetic.mate_1.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None ISF AAAAAAAAAAAAAAAAA XXXXXXXXXXXXXXXXX ../input_files/project1/synthetic.mate_2.fastq.gz AGATCGGAAGAGCGT XXXXXXXXXXXXX XXXXXXXXXXXXXXXXX TTTTTTTTTTTTTTTTT
+synthetic_10_reads_mate_1_synthetic_10_reads_mate_1 se ../input_files/project2/synthetic.mate_1.fastq.gz 75 31 AGATCGGAAGAGCACA XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/annotation.gtf ../input_files/homo_sapiens/genome.fa 100 250 10 EndToEnd None SF AAAAAAAAAAAAAAAAA XXXXXXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX XXXXXXXXXXXXX
diff --git a/tests/input_files/samples_alfa.tsv b/tests/input_files/samples_alfa.tsv
index af1835c28f16f31fe571797e3ae07355d1710e50..9ef1c3cb305934501af14f2752819355081385e5 100644
--- a/tests/input_files/samples_alfa.tsv
+++ b/tests/input_files/samples_alfa.tsv
@@ -1,5 +1,5 @@
-sample seqmode fq1 index_size kmer fq2 fq1_3p fq1_5p fq2_3p fq2_5p organism gtf genome sd mean multimappers soft_clip pass_mode libtype kallisto_directionality fq1_polya fq2_polya alfa_directionality alfa_plus alfa_minus
-paired_end_R1_on_plus_sense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX GATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None A --fr AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT fr-firststrand str1 str2
-paired_end_R1_on_plus_antisense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None A --rf AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT fr-secondstrand str2 str1
-paired_end_R1_on_minus_sense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None A --fr AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT fr-firststrand str1 str2
-paired_end_R1_on_minus_antisense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None A --rf AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT fr-secondstrand str2 str1
+sample seqmode fq1 index_size kmer fq2 fq1_3p fq1_5p fq2_3p fq2_5p organism gtf genome sd mean multimappers soft_clip pass_mode libtype fq1_polya fq2_polya
+paired_end_R1_on_plus_sense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX GATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None ISF AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT
+paired_end_R1_on_plus_antisense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None ISR AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT
+paired_end_R1_on_minus_sense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None ISF AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT
+paired_end_R1_on_minus_antisense pe XXXXXXXXXXXXX 75 31 XXXXXXXXXXXXX AGATCGGAAGAGCACA XXXXXXXXXXXXX AGATCGGAAGAGCGT XXXXXXXXXXXXX homo_sapiens ../input_files/homo_sapiens/quick_start.gtf XXXXXXXXXXXXX 100 250 10 EndToEnd None ISR AAAAAAAAAAAAAAAAA TTTTTTTTTTTTTTTTT
diff --git a/tests/test_scripts_prepare_inputs_table/expected_output.md5 b/tests/test_scripts_prepare_inputs_table/expected_output.md5
index 53afed559996a1e89802ff5bb9307351b8004fdf..aca34c0e6a67951980803d7b7d912ea94ef2d0c1 100644
--- a/tests/test_scripts_prepare_inputs_table/expected_output.md5
+++ b/tests/test_scripts_prepare_inputs_table/expected_output.md5
@@ -1,2 +1,2 @@
40bd0f0fcecdd0d9bc932f63c2811478 config.yaml
-c8dcc5a203e9046806c4090525960151 samples.tsv
+d8fb1773e3b83b6fab0a0d44c9fa71e6 samples.tsv
\ No newline at end of file
diff --git a/workflow/Snakefile b/workflow/Snakefile
index 52317cb1f4e677074ad376a1ad9381db8d71378a..198ae9581a92d27b69aef47e5e312a6af0449e8e 100644
--- a/workflow/Snakefile
+++ b/workflow/Snakefile
@@ -18,6 +18,19 @@ samples_table = pd.read_csv(
sep="\t",
)
+# dict for translation of "directionality parameters"
+directionality_dict = {
+ "SF":
+ {"kallisto":"--fr-stranded",
+ "alfa": "fr-secondstrand",
+ "alfa_plus": "str1",
+ "alfa_minus": "str2"},
+ "SR":
+ {"kallisto":"--rf-stranded",
+ "alfa": "fr-firststrand",
+ "alfa_plus": "str2",
+ "alfa_minus": "str1"},
+}
# Validat config
validate(config, os.path.join("..", "resources", "config_schema.json"))
logger.info(f'Config file after validation: {config}')
@@ -50,6 +63,24 @@ def get_sample(column_id, search_id=None, search_value=None):
else:
return str(samples_table[column_id][0])
+def get_directionality(libtype, tool):
+ """ Get directionality value for different tools"""
+ directionality =""
+
+ for key in directionality_dict.keys():
+ # Use the first of 'SF' or 'SR' that is found in libtype to look up directionality params for the current tool
+ if key in libtype:
+ directionality = directionality_dict[key][tool]
+ break
+
+ # If libtype contains neither 'SF', nor 'SR' we don't know what to do
+ if not directionality:
+ raise ValueError(
+ f"Unknown libtype {libtype}.\n"
+ "Provide one of 'SF', 'SR', 'ISF', 'ISR', 'OSF', 'OSR', 'MSF', 'MSR' in samples.tsv!"
+ )
+
+ return directionality
def parse_rule_config(rule_config: dict, current_rule: str, immutable: Tuple[str, ...] = ()):
"""Get rule specific parameters from rule_config file"""
@@ -1314,10 +1345,10 @@ rule rename_star_rpm_for_alfa:
"{sample}_Signal.{unique}.{plus}.out.bg"),
sample=wildcards.sample,
unique=wildcards.unique,
- plus=get_sample(
- 'alfa_plus',
+ plus=get_directionality(get_sample(
+ 'libtype',
search_id='index',
- search_value=wildcards.sample)),
+ search_value=wildcards.sample),"alfa_plus")),
minus = lambda wildcards:
expand(
os.path.join(
@@ -1328,10 +1359,10 @@ rule rename_star_rpm_for_alfa:
"{sample}_Signal.{unique}.{minus}.out.bg"),
sample=wildcards.sample,
unique=wildcards.unique,
- minus=get_sample(
- 'alfa_minus',
+ minus=get_directionality(get_sample(
+ 'libtype',
search_id='index',
- search_value=wildcards.sample))
+ search_value=wildcards.sample), "alfa_minus"))
output:
plus = temp(os.path.join(
@@ -1514,10 +1545,10 @@ rule alfa_qc:
os.path.basename(input.minus),
name = "{sample}",
alfa_orientation = lambda wildcards:
- get_sample(
- 'alfa_directionality',
- search_id='index',
- search_value=wildcards.sample),
+ get_directionality(get_sample(
+ 'libtype',
+ search_id='index',
+ search_value=wildcards.sample),"alfa"),
additional_params = parse_rule_config(
rule_config,
current_rule=current_rule,
diff --git a/workflow/rules/paired_end.snakefile.smk b/workflow/rules/paired_end.snakefile.smk
index d0a9bc20bb7616a9f41c1bbd7e9ff449062b833d..081fa53fc01ba155f9af6731194626bd5936bbb6 100644
--- a/workflow/rules/paired_end.snakefile.smk
+++ b/workflow/rules/paired_end.snakefile.smk
@@ -496,10 +496,10 @@ rule pe_genome_quantification_kallisto:
"{sample}",
"quant_kallisto"),
directionality = lambda wildcards:
- get_sample(
- 'kallisto_directionality',
- search_id='index',
- search_value=wildcards.sample),
+ get_directionality(get_sample(
+ 'libtype',
+ search_id='index',
+ search_value=wildcards.sample),"kallisto"),
additional_params = parse_rule_config(
rule_config,
current_rule=current_rule,
@@ -536,7 +536,7 @@ rule pe_genome_quantification_kallisto:
-i {input.index} \
-o {params.output_dir} \
-t {threads} \
- {params.directionality}-stranded \
+ {params.directionality} \
{params.additional_params} \
--pseudobam \
{input.reads1} {input.reads2} > {output.pseudoalignment}) \
diff --git a/workflow/rules/single_end.snakefile.smk b/workflow/rules/single_end.snakefile.smk
index 7dfb2c2e2ff38968b61d29fc22773dfe45c82103..c64d71d366cb28897d593e0a714e7579c10e95b0 100644
--- a/workflow/rules/single_end.snakefile.smk
+++ b/workflow/rules/single_end.snakefile.smk
@@ -449,10 +449,10 @@ rule genome_quantification_kallisto:
search_id='index',
search_value=wildcards.sample),
directionality = lambda wildcards:
- get_sample(
- 'kallisto_directionality',
- search_id='index',
- search_value=wildcards.sample),
+ get_directionality(get_sample(
+ 'libtype',
+ search_id='index',
+ search_value=wildcards.sample),"kallisto"),
additional_params = parse_rule_config(
rule_config,
current_rule=current_rule,
@@ -491,7 +491,7 @@ rule genome_quantification_kallisto:
-l {params.fraglen} \
-s {params.fragsd} \
-t {threads} \
- {params.directionality}-stranded \
+ {params.directionality} \
{params.additional_params} \
--pseudobam \
{input.reads} > {output.pseudoalignment};) \