LibMatic pipeline

DIA data processing

cd DIA/
pipeline.pyz pipeline_parameters_DIA.txt program_paths.txt --options 'memory = 100.0 GiB' 'parallelism = 18'

Program arguments:

1. Parameter file
2. File with program paths
3. Memory usage
4. CPU cores to be used

   DDA data processing

   cd DDA/
   pipeline.pyz pipeline_parameters_DDA.txt program_paths.txt --options 'memory = 100.0 GiB' 'parallelism = 18' --DDA
   

   Program arguments: Parameter file

5. File with program paths
6. Memory usage
7. CPU cores to be used
8. Data is DDA

For uncombined DDA/DIA library

LIBGEN_dir=DDA_lib
datadir=./workdir_DDA
fasta_file=./td-up000005640.fasta

For combined (DDA+DIA) library

LIBGEN_dir=DIA_DDA_lib
DIA_datadir=./workdir_DIA
DDA_datadir=./workdir_DDA
fasta_file=./td-up000005640.fasta

Retention time alignment

RTalign.py ${LIBGEN_dir}/RT_align ${datadir}/iproph none /app/teog/tpp5/bin/
RTalign.py ${LIBGEN_dir}/RT_align ${DIA_datadir}/iproph ${DDA_datadir}/iproph /app/teog/tpp5/bin/

Program arguments:

1. Directory to store the RT aligned files
2. Directory of pep.xml files
3. Directory of pep.xml files for DDA (input “none” if not aligning with DDA)
4. Directory of TPP binaries

Combining DIA and DDA pep.xml files

combine_prot_xml.py ${LIBGEN_dir}/combined_prots ${DIA_datadir}/iproph ${DDA_datadir}/iproph /app/teog/tpp5/bin/

Generating spectral library

Run for combined DIA and DDA library

PATH=".:$PATH"
python3 bin/\
gen_con_spec_lib.py \
  ${fasta_file} \
  ${LIBGEN_dir}/RT_align/RT_aligned \
  ${LIBGEN_dir}/combined_prots/interact.prot.xml \
  ${LIBGEN_dir}/DIA_lib true &
python3 bin/\
gen_con_spec_lib.py \
  ${fasta_file} \
  ${LIBGEN_dir}/RT_align/RT_aligned_DDA \
  ${LIBGEN_dir}/combined_prots/interact.prot.xml \
  ${LIBGEN_dir}/DDA_lib true &
wait

For uncombined DIA/DDA library

PATH=".:$PATH" python3 bin/gen_con_spec_lib.py \
  ${fasta_file} \
  ${LIBGEN_dir}/RT_align/RT_aligned \
  ${datadir}/ProteinProphet/interact.prot.xml \
  ${LIBGEN_dir} true

Program arguments:

1. Directory of TPP binaries
2. Path of FASTA file
3. Data directory
4. RT aligned files
5. Output files directory

Combining DIA and DDA generated library

# python3 -m pip -- install matplotlib_venn
(cd ${LIBGEN_dir} && 
python3 ../bin/\
combine_DIA_DDA_libs.py \
  ./DIA_lib/con_lib.tsv \
  ./DDA_lib/con_lib.tsv
)

Program arguments:

1. DIA library
2. DDA library