public virtual List <FeatureLocation> GetSequenceRegions()
{
//Read sequence regions
var items = SequenceRegionUtils.GetSequenceRegions(CoordinateFile);
items.ForEach(m =>
{
if (m.Seqname.StartsWith("chr"))
{
m.Seqname = m.Seqname.StringAfter("chr");
}
});
//Fill sequence information, only miRNA and tRNA will be filled.
if (!string.IsNullOrEmpty(this.FastaFile))
{
Console.WriteLine("Reading sequence from {0} ...", this.FastaFile);
var seqs = SequenceUtils.Read(new FastaFormat(), this.FastaFile).ToDictionary(m => m.Name);
items.ForEach(m =>
{
if (m.Name.StartsWith(SmallRNAConsts.miRNA) || m.Name.StartsWith(SmallRNAConsts.tRNA))
{
if (seqs.ContainsKey(m.Name))
{
m.Sequence = seqs[m.Name].SeqString;
}
else
{
Console.WriteLine("Missing sequence: " + m.Name);
}
}
else
{
m.Sequence = string.Empty;
}
});
seqs.Clear();
}
var result = items.ConvertAll(m => new FeatureLocation(m)).ToList();
result.ForEach(m =>
{
foreach (var categoryName in SmallRNAConsts.Biotypes)
{
if (m.Name.StartsWith(categoryName))
{
m.Category = categoryName;
}
}
});
return(result);
}
public MappedMirnaRegion FindOrCreateRegion(string loc)
{
foreach (var region in MappedRegions)
{
if (region.Region.GetLocation().Equals(loc))
{
return(region);
}
}
var result = new MappedMirnaRegion();
result.Region = SequenceRegionUtils.ParseLocation <SequenceRegion>(loc);
this.MappedRegions.Add(result);
return(result);
}
public virtual List <FeatureLocation> GetSequenceRegions()
{
//Read sequence regions
var result = SequenceRegionUtils.GetSequenceRegions(CoordinateFile, GtfFeatureName, BedAsGtf);
result.ForEach(m =>
{
m.Seqname = m.Seqname.StringAfter("chr");
});
//Fill sequence information
var sr = result.FirstOrDefault(m => m.Name.Contains(":"));
if (sr != null)
{
var sequence = sr.Name.StringAfter(":");
if (sequence.All(m => MIRNA.Contains(m)))
{
result.ForEach(m => m.Sequence = m.Name.StringAfter(":"));
result.ForEach(m => m.Name = m.Name.StringBefore(":"));
}
}
if (!string.IsNullOrEmpty(this.FastaFile))
{
Console.WriteLine("Reading sequence from {0} ...", this.FastaFile);
var seqs = SequenceUtils.Read(new FastaFormat(), this.FastaFile).ToDictionary(m => m.Name);
result.ForEach(m =>
{
if (seqs.ContainsKey(m.Name))
{
m.Sequence = seqs[m.Name].SeqString;
}
else
{
Console.WriteLine("Missing sequence: " + m.Name);
}
});
seqs.Clear();
}
return(result.ConvertAll(m => new FeatureLocation(m)).ToList());
}
public override IEnumerable <string> Process()
{
List <GtfItem> items = new List <GtfItem>();
foreach (var corfile in _options.CoordinateFiles)
{
var curitems = SequenceRegionUtils.GetSequenceRegions(corfile);
if (_options.Features != null && _options.Features.Count > 0)
{
items.RemoveAll(m => !_options.Features.Contains(m.Feature));
}
if (!string.IsNullOrEmpty(_options.NameKey))
{
var key = _options.NameKey + "=";
items.ForEach(m =>
{
if (!string.IsNullOrEmpty(m.Attributes) && m.Attributes.Contains(key))
{
m.Name = m.Attributes.StringAfter(key).StringBefore(";").Trim();
}
});
}
items.AddRange(curitems);
}
var removechr = !items.All(m => m.Seqname.StartsWith("chr"));
if (removechr)
{
items.ForEach(m => m.Seqname = m.Seqname.StringAfter("chr"));
}
var map = items.GroupBy(m => m.Seqname).ToDictionary(m => m.Key, m => m.ToList());
using (StreamWriter sw = new StreamWriter(_options.OutputFile))
{
using (var sr = new StreamReader(_options.InputFile))
{
string line;
var headers = sr.ReadLine().Split(',').ToList();
headers.Add("ClosetFeature");
headers.Add("ClosetFeatureLocus");
headers.Add("ClosetFeatureDistance");
sw.WriteLine(headers.Merge(','));
List <GtfItem> mingtfs = new List <GtfItem>();
while ((line = sr.ReadLine()) != null)
{
var parts = line.Split(',');
var chr = parts[0];
if (removechr)
{
chr = chr.StringAfter("chr");
}
var start = long.Parse(parts[2]);
var end = long.Parse(parts[3]);
var sequence = parts[5];
var location = int.Parse(parts[7]);
var t2c_feature = string.Empty;
List <GtfItem> gtfs;
if (!map.TryGetValue(chr, out gtfs))
{
sw.WriteLine("{0},,,", line);
continue;
}
long mindist = int.MaxValue;
mingtfs.Clear();
foreach (var gtf in gtfs)
{
long dist;
if (gtf.Start > end)
{
dist = gtf.Start - end;
}
else if (gtf.End < start)
{
dist = start - gtf.End;
}
else
{
dist = 0;
}
if (dist < mindist)
{
mingtfs.Clear();
mingtfs.Add(gtf);
mindist = dist;
}
else if (dist == mindist)
{
mingtfs.Add(gtf);
}
}
sw.WriteLine("{0},{1},{2},{3}",
line,
(from m in mingtfs select m.Name).Merge(";"),
(from m in mingtfs select m.GetLocation()).Merge(";"),
mindist);
}
}
}
return(new string[] { _options.OutputFile });
}
public override IEnumerable <string> Process()
{
PileupCountList pc = new PileupCountList();
var format = options.GetSAMFormat();
var cm = new SmallRNACountMap(options.CountFile);
var srItems = SequenceRegionUtils.GetSequenceRegions(options.CoordinateFile, "miRNA", options.BedAsGtf);
srItems.ForEach(m =>
{
m.Seqname = m.Seqname.StringAfter("chr");
});
var srmap = srItems.GroupBy(m => m.Seqname).ToDictionary(m => m.Key, m => m.ToList());
StreamWriter swScript = null;
try
{
if (options.ExportIgvScript)
{
swScript = new StreamWriter(options.OutputFile + ".igv");
swScript.WriteLine("snapshotDirectory {0}", Path.GetDirectoryName(options.OutputFile).Replace('\\', '/'));
}
using (StreamWriter sw = new StreamWriter(options.OutputFile))
{
sw.WriteLine(@"##fileformat=VCFv4.2
##fileDate={0:yyyyMMdd}
##source={1}
##phasing=partial
##INFO=<ID=NS,Number=1,Type=Integer,Description=""Number of Samples With Data"">
##INFO=<ID=DP,Number=1,Type=Integer,Description=""Total Depth"">
##INFO=<ID=AF,Number=A,Type=Float,Description=""Allele Frequency"">
##INFO=<ID=FP,Number=1,Type=Float,Description=""Fisher Exact Test P-Value"">
##INFO=<ID=MN,Number=.,Type=String,Description=""miRNA name contains this position"">
##FILTER=<ID=FisherET,Description=""Fisher exact test Pvalue less than {2}"">
##FILTER=<ID=AltAlleFreq,Description=""Alternative allele frequency less than {3}"">
##FILTER=<ID=notMiRNA,Description=""Position not located in miRNA locus"">
##FORMAT=<ID=DP,Number=1,Type=Integer,Description=""Read Depth"">
##FORMAT=<ID=AD,Number=1,Type=Integer,Description=""Allelic Depth"">
#CHROM POS ID REF ALT QUAL FILTER INFO FORMAT {4}",
DateTime.Now,
"PileupCountBuilder",
options.FisherPValue,
options.MinimumAlternativeAlleleFrequency,
Path.GetFileNameWithoutExtension(options.InputFile));
using (var sr = SAMFactory.GetReader(options.InputFile, true))
{
int count = 0;
string line;
while ((line = sr.ReadLine()) != null)
{
count++;
if (count % 100 == 0)
{
if (Progress.IsCancellationPending())
{
throw new UserTerminatedException();
}
}
if (count % 100000 == 0)
{
Progress.SetMessage("{0} reads processed", count);
}
var parts = line.Split('\t');
var qname = parts[SAMFormatConst.QNAME_INDEX];
var seq = parts[SAMFormatConst.SEQ_INDEX];
//too short
if (seq.Length < options.MinimumReadLength)
{
continue;
}
SAMFlags flag = (SAMFlags)int.Parse(parts[SAMFormatConst.FLAG_INDEX]);
//unmatched
if (flag.HasFlag(SAMFlags.UnmappedQuery))
{
continue;
}
var cigar = parts[SAMFormatConst.CIGAR_INDEX];
//insertion/deletion
if (cigar.Any(m => m == 'I' || m == 'D'))
{
continue;
}
var sam = new SAMAlignedItem()
{
Qname = qname,
};
bool isReversed = flag.HasFlag(SAMFlags.QueryOnReverseStrand);
char strand;
if (isReversed)
{
strand = '-';
sam.Sequence = SequenceUtils.GetReverseComplementedSequence(seq);
}
else
{
strand = '+';
sam.Sequence = seq;
}
var loc = new SAMAlignedLocation(sam)
{
Seqname = parts[SAMFormatConst.RNAME_INDEX].StringAfter("chr"),
Start = int.Parse(parts[SAMFormatConst.POS_INDEX]),
Strand = strand,
Cigar = parts[SAMFormatConst.CIGAR_INDEX],
MismatchPositions = format.GetMismatchPositions(parts),
NumberOfMismatch = format.GetNumberOfMismatch(parts),
Sequence = seq
};
loc.ParseEnd(sam.Sequence);
sam.AddLocation(loc);
if (format.HasAlternativeHits)
{
format.ParseAlternativeHits(parts, sam);
}
var finished = pc.Add(sam, cm.GetCount(sam.Qname));
if (null == finished || 0 == finished.Count)
{
continue;
}
foreach (var fin in finished)
{
//if (fin.Chromosome.Equals("1") && fin.Position == 5160725)
//{
// Console.WriteLine(fin);
//}
var ft = fin.FisherExactTest();
if (ft.PValue <= options.FisherPValue)
{
var total = fin.Sum(m => m.Value);
var minallele = total * options.MinimumAlternativeAlleleFrequency;
if (ft.Sample2.Failed >= minallele)
{
List <GtfItem> srs;
List <string> ranges = new List <string>();
if (srmap.TryGetValue(sam.Locations[0].Seqname, out srs))
{
foreach (var seqr in srs)
{
if (seqr.Contains(fin.Position))
{
ranges.Add(seqr.GetNameLocation());
}
}
}
var alter = (from r in fin
where r.Key != fin.Reference
orderby r.Key
select r).ToList();
var str = string.Format("{0}\t{1}\t{2}\t{3}\t{4}\t{5}\t{6}\tNS={7};DP={8};AF={9};FP={10:0.##E0}{11}\tDP:AD\t{12}:{13},{14}",
fin.Chromosome,
fin.Position,
".",
fin.Reference,
(from r in alter
select r.Key.ToString()).Merge(","),
0,
ranges.Count == 0 ? "notMiRNA" : "PASS",
1,
total,
(from r in alter
select string.Format("{0:0.###}", r.Value * 1.0 / total)).Merge(","),
ft.PValue,
ranges.Count == 0 ? "" : ";" + ranges.Merge(","),
total,
ft.Sample2.Succeed,
(from r in alter
select r.Value.ToString()).Merge(","));
sw.WriteLine(str);
//Console.WriteLine(str);
if (swScript != null && ranges.Count > 0)
{
swScript.WriteLine(@"goto {0}:{1}
sort position
snapshot {0}_{2}_{1}.png", fin.Chromosome, fin.Position, ranges[0].Replace('(', '_').Replace(')', '_').Replace(':', '_'));
}
}
}
}
finished.Clear();
}
}
}
}
finally
{
if (swScript != null)
{
swScript.Close();
}
}
return(new string[] { options.OutputFile });
}
public override IEnumerable <string> Process()
{
var srItems = SequenceRegionUtils.GetSequenceRegions(options.InputFile).Where(m => options.AcceptName(m.Name)).ToList();
srItems = (from sr in srItems.GroupBy(m => m.Name)
select sr.First()).ToList();
var keepChrInName = options.KeepChrInName && srItems.Any(m => m.Name.StartsWith("chr"));
if (!keepChrInName)
{
srItems.ForEach(m => m.Seqname = m.Seqname.StringAfter("chr"));
}
var srMap = srItems.ToGroupDictionary(m => m.Seqname);
var ff = new FastaFormat(int.MaxValue);
using (StreamWriter sw = new StreamWriter(options.OutputFile))
{
using (StreamReader sr = new StreamReader(options.GenomeFastaFile))
{
Sequence seq;
while ((seq = ff.ReadSequence(sr)) != null)
{
Progress.SetMessage("processing " + seq.Name + " ...");
var name = seq.Name;
if (!keepChrInName)
{
name = name.StringAfter("chr");
}
List <GtfItem> items;
if (!srMap.TryGetValue(name, out items))
{
if (name.Equals("M"))
{
name = "MT";
srMap.TryGetValue(name, out items);
}
else if (name.Equals("chrM"))
{
name = "chrMT";
srMap.TryGetValue(name, out items);
}
else if (name.Equals("MT"))
{
name = "M";
srMap.TryGetValue(name, out items);
}
else if (name.Equals("chrMT"))
{
name = "chrM";
srMap.TryGetValue(name, out items);
}
}
if (items != null)
{
Progress.SetMessage(" there are {0} entries in {1} ...", items.Count, name);
foreach (var item in items)
{
if (item.Start - 1 + item.Length >= seq.SeqString.Length)
{
throw new Exception(string.Format("{0} exceed chromosome {1} length {2}", item, name, seq.SeqString.Length));
}
var newseq = seq.SeqString.Substring((int)item.Start - 1, (int)item.Length);
if (item.Strand == '-')
{
newseq = SequenceUtils.GetReverseComplementedSequence(newseq);
}
newseq = newseq.ToUpper();
var newname = string.Format("{0} {1} {2}", item.Name, item.GetLocationWithoutStrand(), item.Strand);
var entry = new Sequence(newname, newseq);
ff.WriteSequence(sw, entry);
}
}
}
}
}
return(new string[] { options.OutputFile });
}
