ExitCodes main_(int, const char**)
{
String inputfile_name = getStringOption_("in");
String outputfile_name = getStringOption_("out");
vector<ProteinIdentification> proteins;
vector<PeptideIdentification> peptides;
IdXMLFile().load(inputfile_name, proteins, peptides);
Size n_prot_ids = proteins.size();
Size n_prot_hits = IDFilter::countHits(proteins);
Size n_pep_ids = peptides.size();
Size n_pep_hits = IDFilter::countHits(peptides);
// Filtering peptide identification according to set criteria
double rt_high = numeric_limits<double>::infinity(), rt_low = -rt_high;
if (parseRange_(getStringOption_("precursor:rt"), rt_low, rt_high))
{
LOG_INFO << "Filtering peptide IDs by precursor RT..." << endl;
IDFilter::filterPeptidesByRT(peptides, rt_low, rt_high);
}
double mz_high = numeric_limits<double>::infinity(), mz_low = -mz_high;
if (parseRange_(getStringOption_("precursor:mz"), mz_low, mz_high))
{
LOG_INFO << "Filtering peptide IDs by precursor m/z...";
IDFilter::filterPeptidesByMZ(peptides, mz_low, mz_high);
}
// Filtering peptide hits according to set criteria
if (getFlag_("unique"))
{
LOG_INFO << "Removing duplicate peptide hits..." << endl;
IDFilter::removeDuplicatePeptideHits(peptides);
}
if (getFlag_("unique_per_protein"))
{
LOG_INFO << "Filtering peptides by unique match to a protein..." << endl;
IDFilter::keepUniquePeptidesPerProtein(peptides);
}
double peptide_significance = getDoubleOption_("thresh:pep");
if (peptide_significance > 0)
{
LOG_INFO << "Filtering by peptide significance threshold..." << endl;
IDFilter::filterHitsBySignificance(peptides, peptide_significance);
}
double pred_rt_pv = getDoubleOption_("rt:p_value");
if (pred_rt_pv > 0)
{
LOG_INFO << "Filtering by RT prediction p-value..." << endl;
IDFilter::filterPeptidesByRTPredictPValue(
peptides, "predicted_RT_p_value", pred_rt_pv);
}
double pred_rt_pv_1d = getDoubleOption_("rt:p_value_1st_dim");
if (pred_rt_pv_1d > 0)
{
LOG_INFO << "Filtering by RT prediction p-value (first dim.)..." << endl;
IDFilter::filterPeptidesByRTPredictPValue(
peptides, "predicted_RT_p_value_first_dim", pred_rt_pv_1d);
}
String whitelist_fasta = getStringOption_("whitelist:proteins").trim();
if (!whitelist_fasta.empty())
{
LOG_INFO << "Filtering by protein whitelisting (FASTA input)..." << endl;
// load protein accessions from FASTA file:
vector<FASTAFile::FASTAEntry> fasta;
FASTAFile().load(whitelist_fasta, fasta);
set<String> accessions;
for (vector<FASTAFile::FASTAEntry>::iterator it = fasta.begin();
it != fasta.end(); ++it)
{
accessions.insert(it->identifier);
}
IDFilter::keepHitsMatchingProteins(peptides, accessions);
IDFilter::keepHitsMatchingProteins(proteins, accessions);
}
vector<String> whitelist_accessions =
getStringList_("whitelist:protein_accessions");
if (!whitelist_accessions.empty())
{
LOG_INFO << "Filtering by protein whitelisting (accessions input)..."
<< endl;
set<String> accessions(whitelist_accessions.begin(),
whitelist_accessions.end());
IDFilter::keepHitsMatchingProteins(peptides, accessions);
IDFilter::keepHitsMatchingProteins(proteins, accessions);
}
String whitelist_peptides = getStringOption_("whitelist:peptides").trim();
if (!whitelist_peptides.empty())
{
LOG_INFO << "Filtering by inclusion peptide whitelisting..." << endl;
vector<PeptideIdentification> inclusion_peptides;
vector<ProteinIdentification> inclusion_proteins; // ignored
IdXMLFile().load(whitelist_peptides, inclusion_proteins,
inclusion_peptides);
bool ignore_mods = getFlag_("whitelist:ignore_modifications");
IDFilter::keepPeptidesWithMatchingSequences(peptides, inclusion_peptides,
ignore_mods);
}
vector<String> whitelist_mods = getStringList_("whitelist:modifications");
if (!whitelist_mods.empty())
{
LOG_INFO << "Filtering peptide IDs by modification whitelisting..."
<< endl;
set<String> good_mods(whitelist_mods.begin(), whitelist_mods.end());
IDFilter::keepPeptidesWithMatchingModifications(peptides, good_mods);
}
String blacklist_fasta = getStringOption_("blacklist:proteins").trim();
if (!blacklist_fasta.empty())
{
LOG_INFO << "Filtering by protein blacklisting (FASTA input)..." << endl;
// load protein accessions from FASTA file:
vector<FASTAFile::FASTAEntry> fasta;
FASTAFile().load(blacklist_fasta, fasta);
set<String> accessions;
for (vector<FASTAFile::FASTAEntry>::iterator it = fasta.begin();
it != fasta.end(); ++it)
{
accessions.insert(it->identifier);
}
IDFilter::removeHitsMatchingProteins(peptides, accessions);
IDFilter::removeHitsMatchingProteins(proteins, accessions);
}
vector<String> blacklist_accessions =
getStringList_("blacklist:protein_accessions");
if (!blacklist_accessions.empty())
{
LOG_INFO << "Filtering by protein blacklisting (accessions input)..."
<< endl;
set<String> accessions(blacklist_accessions.begin(),
blacklist_accessions.end());
IDFilter::removeHitsMatchingProteins(peptides, accessions);
IDFilter::removeHitsMatchingProteins(proteins, accessions);
}
String blacklist_peptides = getStringOption_("blacklist:peptides").trim();
if (!blacklist_peptides.empty())
{
LOG_INFO << "Filtering by exclusion peptide blacklisting..." << endl;
vector<PeptideIdentification> exclusion_peptides;
vector<ProteinIdentification> exclusion_proteins; // ignored
IdXMLFile().load(blacklist_peptides, exclusion_proteins,
exclusion_peptides);
bool ignore_mods = getFlag_("blacklist:ignore_modifications");
IDFilter::removePeptidesWithMatchingSequences(
peptides, exclusion_peptides, ignore_mods);
}
vector<String> blacklist_mods = getStringList_("blacklist:modifications");
if (!blacklist_mods.empty())
{
LOG_INFO << "Filtering peptide IDs by modification blacklisting..."
<< endl;
set<String> bad_mods(blacklist_mods.begin(), blacklist_mods.end());
IDFilter::removePeptidesWithMatchingModifications(peptides, bad_mods);
}
if (getFlag_("best:strict"))
{
LOG_INFO << "Filtering by best peptide hits..." << endl;
IDFilter::keepBestPeptideHits(peptides, true);
}
Int min_length = 0, max_length = 0;
if (parseRange_(getStringOption_("length"), min_length, max_length))
{
LOG_INFO << "Filtering by peptide length..." << endl;
if ((min_length < 0) || (max_length < 0))
{
LOG_ERROR << "Fatal error: negative values are not allowed for parameter 'length'" << endl;
return ILLEGAL_PARAMETERS;
}
IDFilter::filterPeptidesByLength(peptides, Size(min_length),
Size(max_length));
}
// Filter by digestion enzyme product
String protein_fasta = getStringOption_("digest:fasta").trim();
if (!protein_fasta.empty())
{
LOG_INFO << "Filtering peptides by digested protein (FASTA input)..." << endl;
// load protein accessions from FASTA file:
vector<FASTAFile::FASTAEntry> fasta;
FASTAFile().load(protein_fasta, fasta);
// Configure Enzymatic digestion
EnzymaticDigestion digestion;
String enzyme = getStringOption_("digest:enzyme").trim();
if (!enzyme.empty())
{
digestion.setEnzyme(enzyme);
}
String specificity = getStringOption_("digest:specificity").trim();
if (!specificity.empty())
{
digestion.setSpecificity(digestion.getSpecificityByName(specificity));
}
Int missed_cleavages = getIntOption_("digest:missed_cleavages");
bool ignore_missed_cleavages = true;
if (missed_cleavages > -1)
{
ignore_missed_cleavages = false;
if (digestion.getSpecificity() == EnzymaticDigestion::SPEC_FULL)
{
LOG_WARN << "Specificity not full, missed_cleavages option is redundant" << endl;
}
digestion.setMissedCleavages(missed_cleavages);
}
bool methionine_cleavage = false;
if (getFlag_("digest:methionine_cleavage"))
{
methionine_cleavage = true;
}
// Build the digest filter function
IDFilter::DigestionFilter filter(fasta,
digestion,
ignore_missed_cleavages,
methionine_cleavage);
// Filter peptides
filter.filterPeptideEvidences(peptides);
}
if (getFlag_("var_mods"))
{
LOG_INFO << "Filtering for variable modifications..." << endl;
// gather possible variable modifications from search parameters:
set<String> var_mods;
for (vector<ProteinIdentification>::iterator prot_it = proteins.begin();
prot_it != proteins.end(); ++prot_it)
{
const ProteinIdentification::SearchParameters& params =
prot_it->getSearchParameters();
for (vector<String>::const_iterator mod_it =
params.variable_modifications.begin(); mod_it !=
params.variable_modifications.end(); ++mod_it)
{
var_mods.insert(*mod_it);
}
}
IDFilter::keepPeptidesWithMatchingModifications(peptides, var_mods);
}
double pep_score = getDoubleOption_("score:pep");
// @TODO: what if 0 is a reasonable cut-off for some score?
if (pep_score != 0)
{
LOG_INFO << "Filtering by peptide score..." << endl;
IDFilter::filterHitsByScore(peptides, pep_score);
}
Int min_charge = numeric_limits<Int>::min(), max_charge =
numeric_limits<Int>::max();
if (parseRange_(getStringOption_("charge"), min_charge, max_charge))
{
LOG_INFO << "Filtering by peptide charge..." << endl;
IDFilter::filterPeptidesByCharge(peptides, min_charge, max_charge);
}
Size best_n_pep = getIntOption_("best:n_peptide_hits");
if (best_n_pep > 0)
{
LOG_INFO << "Filtering by best n peptide hits..." << endl;
IDFilter::keepNBestHits(peptides, best_n_pep);
}
Int min_rank = 0, max_rank = 0;
if (parseRange_(getStringOption_("best:n_to_m_peptide_hits"), min_rank,
max_rank))
{
LOG_INFO << "Filtering by peptide hit ranks..." << endl;
if ((min_rank < 0) || (max_rank < 0))
{
LOG_ERROR << "Fatal error: negative values are not allowed for parameter 'best:n_to_m_peptide_hits'" << endl;
return ILLEGAL_PARAMETERS;
}
IDFilter::filterHitsByRank(peptides, Size(min_rank), Size(max_rank));
}
double mz_error = getDoubleOption_("mz:error");
if (mz_error > 0)
{
LOG_INFO << "Filtering by mass error..." << endl;
bool unit_ppm = (getStringOption_("mz:unit") == "ppm");
IDFilter::filterPeptidesByMZError(peptides, mz_error, unit_ppm);
}
// Filtering protein identifications according to set criteria
double protein_significance = getDoubleOption_("thresh:prot");
if (protein_significance > 0)
{
LOG_INFO << "Filtering by protein significance threshold..." << endl;
IDFilter::filterHitsBySignificance(proteins, protein_significance);
}
double prot_score = getDoubleOption_("score:prot");
// @TODO: what if 0 is a reasonable cut-off for some score?
if (prot_score != 0)
{
LOG_INFO << "Filtering by protein score..." << endl;
IDFilter::filterHitsByScore(proteins, prot_score);
}
Size best_n_prot = getIntOption_("best:n_protein_hits");
if (best_n_prot > 0)
{
LOG_INFO << "Filtering by best n protein hits..." << endl;
IDFilter::keepNBestHits(proteins, best_n_prot);
}
if (getFlag_("remove_decoys"))
{
LOG_INFO << "Removing decoy hits..." << endl;
IDFilter::removeDecoyHits(peptides);
IDFilter::removeDecoyHits(proteins);
}
// Clean-up:
if (!getFlag_("keep_unreferenced_protein_hits"))
{
LOG_INFO << "Removing unreferenced protein hits..." << endl;
IDFilter::removeUnreferencedProteins(proteins, peptides);
}
IDFilter::updateHitRanks(proteins);
IDFilter::updateHitRanks(peptides);
// remove non-existant protein references from peptides (and optionally:
// remove peptides with no proteins):
bool rm_pep = getFlag_("delete_unreferenced_peptide_hits");
if (rm_pep) LOG_INFO << "Removing peptide hits without protein references..." << endl;
IDFilter::updateProteinReferences(peptides, proteins, rm_pep);
IDFilter::removeEmptyIdentifications(peptides);
// we want to keep "empty" protein IDs because they contain search meta data
// update protein groupings if necessary:
for (vector<ProteinIdentification>::iterator prot_it = proteins.begin();
prot_it != proteins.end(); ++prot_it)
{
bool valid = IDFilter::updateProteinGroups(prot_it->getProteinGroups(),
prot_it->getHits());
if (!valid)
{
LOG_WARN << "Warning: While updating protein groups, some proteins were removed from groups that are still present. The new grouping (especially the group probabilities) may not be completely valid any more." << endl;
}
valid = IDFilter::updateProteinGroups(
prot_it->getIndistinguishableProteins(), prot_it->getHits());
if (!valid)
{
LOG_WARN << "Warning: While updating indistinguishable proteins, some proteins were removed from groups that are still present. The new grouping (especially the group probabilities) may not be completely valid any more." << endl;
}
}
// some stats
LOG_INFO << "Before filtering:\n"
<< n_prot_ids << " protein identification(s) with "
<< n_prot_hits << " protein hit(s),\n"
<< n_pep_ids << " peptide identification(s) with "
<< n_pep_hits << " peptides hit(s).\n"
<< "After filtering:\n"
<< proteins.size() << " protein identification(s) with "
<< IDFilter::countHits(proteins) << " protein hit(s),\n"
<< peptides.size() << " peptide identification(s) with "
<< IDFilter::countHits(peptides) << " peptides hit(s)." << endl;
IdXMLFile().store(outputfile_name, proteins, peptides);
return EXECUTION_OK;
}
START_SECTION([EXTRA] ~EnzymaticDigestion())
delete e_ptr;
END_SECTION
START_SECTION((EnzymaticDigestion(const EnzymaticDigestion &rhs)))
EnzymaticDigestion ed;
ed.setMissedCleavages(1234);
ed.setEnzyme("no cleavage");
ed.setSpecificity(EnzymaticDigestion::SPEC_SEMI);
EnzymaticDigestion ed2(ed);
TEST_EQUAL(ed.getMissedCleavages(), ed2.getMissedCleavages());
TEST_EQUAL(ed.getEnzymeName(), ed2.getEnzymeName());
TEST_EQUAL(ed.getSpecificity(), ed2.getSpecificity());
END_SECTION
START_SECTION((EnzymaticDigestion & operator=(const EnzymaticDigestion &rhs)))
EnzymaticDigestion ed;
ed.setMissedCleavages(1234);
ed.setEnzyme("no cleavage");
ed.setSpecificity(EnzymaticDigestion::SPEC_SEMI);
EnzymaticDigestion ed2 = ed;
TEST_EQUAL(ed.getMissedCleavages(), ed2.getMissedCleavages());
TEST_EQUAL(ed.getEnzymeName(), ed2.getEnzymeName());
TEST_EQUAL(ed.getSpecificity(), ed2.getSpecificity());