/// <summary>
/// Converts the Sequence to a QualitativeSequence in the alignment.
/// </summary>
/// <param name="aln">Aln.</param>
/// <param name="qualScores">Qual scores.</param>
public static void ConvertAlignedSequenceToQualSeq(IPairwiseSequenceAlignment aln, int[] qualScores) {
var q = aln.PairwiseAlignedSequences [0].SecondSequence as Sequence;
var qvs = new int[q.Count];
int queryPos = 0;
for (int i = 0; i < qvs.Length; i++) {
if (q [i] == '-') {
qvs [i] = 0;
} else {
qvs [i] = qualScores[queryPos++];
}
}
var qseq = new QualitativeSequence (DnaAlphabet.Instance, FastQFormatType.Sanger, q.ToArray (), qvs, false);
aln.PairwiseAlignedSequences [0].SecondSequence = qseq;
}
public void TestConstructorWithByteArray()
{
byte[] sequenceData = new byte[6];
sequenceData[0] = (byte)'C';
sequenceData[1] = (byte)'A';
sequenceData[2] = (byte)'A';
sequenceData[3] = (byte)'G';
sequenceData[4] = (byte)'C';
sequenceData[5] = (byte)'T';
byte[] qualityScores = new byte[6];
qualityScores[0] = 65;
qualityScores[1] = 65;
qualityScores[2] = 65;
qualityScores[3] = 65;
qualityScores[4] = 110;
qualityScores[5] = 125;
string expectedSequence = "CAAGCT";
QualitativeSequence qualitativeSequence = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Illumina_v1_3, sequenceData, qualityScores);
string actual = "";
foreach (byte bt in qualitativeSequence)
{
actual += (char)bt;
}
Assert.AreEqual(expectedSequence, actual);
Assert.AreEqual(qualitativeSequence.Alphabet, Alphabets.DNA);
Assert.AreEqual(qualitativeSequence.Count, 6);
//
// Test for indexer
Assert.AreEqual(qualitativeSequence[0], (byte)'C');
Assert.AreEqual(qualitativeSequence[1], (byte)'A');
Assert.AreEqual(qualitativeSequence[2], (byte)'A');
Assert.AreEqual(qualitativeSequence[3], (byte)'G');
Assert.AreEqual(qualitativeSequence[4], (byte)'C');
Assert.AreEqual(qualitativeSequence[5], (byte)'T');
int index = 0;
foreach (byte qualityScore in qualitativeSequence.GetEncodedQualityScores())
{
Assert.AreEqual(qualityScores[index++], qualityScore);
}
}
/// <summary>
/// Convert quality values to string array
/// </summary>
/// <param name="sequence">Sequence which has the quality values</param>
/// <param name="maxColumns">Max number of columns to write to</param>
/// <returns>string array with quality values</returns>
public static string[,] FastQQualityValuesToRange(QualitativeSequence sequence, int maxColumns)
{
var qualityScoreArray = sequence.GetEncodedQualityScores();
long rowCount = (int)Math.Ceiling((decimal)qualityScoreArray.Length / (decimal)maxColumns);
long columnCount = sequence.Count > maxColumns ? maxColumns : sequence.Count;
string[,] rangeData = new string[rowCount, columnCount];
int count = 0;
for (int row = 0; row < rowCount; row++)
{
for (int col = 0; col < columnCount && count < qualityScoreArray.Length; col++, count++)
{
rangeData[row, col] = (qualityScoreArray[count]).ToString(CultureInfo.InvariantCulture);
}
}
return rangeData;
}
/// <summary>
/// Initializes a new instance of the <see cref="Bio.IO.PacBio.PacBioCCSRead"/> class. From an initially parsed BAM file.
/// </summary>
/// <param name="s">S.</param>
public PacBioCCSRead (SAMAlignedSequence s)
{
/* TODO: Converting from binary to string and back is beyond silly...
* no performance hit worth worrying about at present, but in the future it might be worth
* going directly from binary to the type rather than through string intermediates */
foreach (var v in s.OptionalFields) {
if (v.Tag == "sn") {
var snrs = v.Value.Split (',').Skip (1).Select (x => Convert.ToSingle (x)).ToArray ();
SnrA = snrs [0];
SnrC = snrs [1];
SnrG = snrs [2];
SnrT = snrs [3];
} else if (v.Tag == "zm") {
HoleNumber = (int)Convert.ToInt32 (v.Value);
} else if (v.Tag == "pq") {
// This tag is now deprecated by the rq tag
ReadQuality = Convert.ToSingle (v.Value);
} else if (v.Tag == "rq") {
ReadQuality = Convert.ToSingle (v.Value);
}else if (v.Tag == "za") {
AvgZscore = (float)Convert.ToSingle (v.Value);
} else if (v.Tag == "rs") {
statusCounts = v.Value.Split (',').Skip (1).Select (x => Convert.ToInt32 (x)).ToArray ();
} else if (v.Tag == "np") {
NumPasses = Convert.ToInt32 (v.Value);
} else if (v.Tag == "RG") {
ReadGroup = v.Value;
} else if (v.Tag == "zs") {
ZScores = v.Value.Split (',').Skip (1).Select (x => Convert.ToSingle (x)).ToArray ();
}
}
// TODO: We should use String.Intern here, but not available in PCL...
// Movie = String.Intern(s.QuerySequence.ID.Split ('/') [0]);
Movie = s.QuerySequence.ID.Split ('/') [0];
Sequence = s.QuerySequence as QualitativeSequence;
}
public void TestGetPhredQualityScore()
{
// Validate using SangerFormat.
List<int> pharedQualityScores = GetPharedQualityScoresForSanger();
byte[] encodedSangerQualityScores = GetSangerEncodedQualityScores(pharedQualityScores);
byte[] symbols = GetSymbols(encodedSangerQualityScores.Length);
QualitativeSequence qualSeq = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Sanger, symbols, encodedSangerQualityScores);
for (int i = 0; i < qualSeq.Count; i++)
{
Assert.AreEqual(pharedQualityScores[i], qualSeq.GetPhredQualityScore(i));
}
// Validate using illumina v1.3 .
pharedQualityScores = GetPharedQualityScoresForIllumina_v1_3();
byte[] encodedIllumina_v1_3_QualityScores = GetIllumina_v1_3_EncodedQualityScores(pharedQualityScores);
symbols = GetSymbols(encodedIllumina_v1_3_QualityScores.Length);
qualSeq = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Illumina_v1_3, symbols, encodedIllumina_v1_3_QualityScores);
for (int i = 0; i < qualSeq.Count; i++)
{
Assert.AreEqual(pharedQualityScores[i], qualSeq.GetPhredQualityScore(i));
}
// Validate using illumina v1.5
pharedQualityScores = GetPharedQualityScoresForIllumina_v1_5();
byte[] encodedIllumina_v1_5_QualityScores = GetIllumina_v1_5_EncodedQualityScores(pharedQualityScores);
symbols = GetSymbols(encodedIllumina_v1_5_QualityScores.Length);
qualSeq = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Illumina_v1_5, symbols, encodedIllumina_v1_5_QualityScores);
for (int i = 0; i < qualSeq.Count; i++)
{
Assert.AreEqual(pharedQualityScores[i], qualSeq.GetPhredQualityScore(i));
}
// Validate using illumina v1.8
pharedQualityScores = GetPharedQualityScoresForIllumina_v1_8();
byte[] encodedIllumina_v1_8_QualityScores = GetIllumina_v1_8_EncodedQualityScores(pharedQualityScores);
symbols = GetSymbols(encodedIllumina_v1_8_QualityScores.Length);
qualSeq = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Illumina_v1_8, symbols, encodedIllumina_v1_8_QualityScores);
for (int i = 0; i < qualSeq.Count; i++)
{
Assert.AreEqual(pharedQualityScores[i], qualSeq.GetPhredQualityScore(i));
}
// Validate using illumina v1.0 .
List<int> solexaQualityScores = GetSolexaQualityScoresForIllumina_v1_0();
byte[] encodedIllumina_v1_0_QualityScores = GetIllumina_v1_0_EncodedQualityScores(solexaQualityScores);
symbols = GetSymbols(encodedIllumina_v1_0_QualityScores.Length);
qualSeq = new QualitativeSequence(Alphabets.DNA, FastQFormatType.Solexa_Illumina_v1_0, symbols, encodedIllumina_v1_0_QualityScores);
for (int i = 0; i < qualSeq.Count; i++)
{
Assert.AreEqual(solexaQualityScores[i], qualSeq.GetSolexaQualityScore(i));
}
}
/// <summary>
/// Given two byte arrays representing a pairwise alignment, shift them so
/// that all deletions start as early as possible. For example:
///
/// <code>
/// TTTTAAAATTTT -> Converts to -> TTTTAAAATTTT
/// TTTTAA--TTTT TTTT--AATTTT
/// </code>
///
/// This function takes a IPairwiseSequenceAlignment and assumes that the first sequence is the reference and second
/// sequence is the query. It returns a new Pairwise sequence alignment with all of the indels left aligned as well as a list of variants.
/// </summary>
/// <param name="aln">Aln. The second sequence should be of type QualitativeSequence or Sequence</param>
/// <param name="callVariants">callVariants. If true, it will call variants, otherwise the second half of tuple will be null. </param>
public static Tuple<IPairwiseSequenceAlignment, List<Variant>> LeftAlignIndelsAndCallVariants(IPairwiseSequenceAlignment aln, bool callVariants = true) {
if (aln == null) {
throw new NullReferenceException ("aln");
}
if (aln.PairwiseAlignedSequences == null || aln.PairwiseAlignedSequences.Count != 1) {
throw new ArgumentException ("The pairwise aligned sequence should only have one alignment");
}
var frstAln = aln.PairwiseAlignedSequences.First ();
var seq1 = frstAln.FirstSequence;
var seq2 = frstAln.SecondSequence;
if (seq1 == null) {
throw new NullReferenceException ("seq1");
} else if (seq2 == null) {
throw new NullReferenceException ("seq2");
}
//TODO: Might implement an ambiguity check later.
#if FALSE
if (seq1.Alphabet.HasAmbiguity || seq2.Alphabet.HasAmbiguity) {
throw new ArgumentException ("Cannot left align sequences with ambiguous symbols.");
}
#endif
// Note we have to copy unless we can guarantee the array will not be mutated.
byte[] refseq = seq1.ToArray ();
ISequence newQuery;
List<Variant> variants = null;
// Call variants for a qualitative sequence
if (seq2 is QualitativeSequence) {
var qs = seq2 as QualitativeSequence;
var query = Enumerable.Zip (qs, qs.GetQualityScores (), (bp, qv) => new BPandQV (bp, (byte)qv, false)).ToArray ();
AlignmentUtils.LeftAlignIndels (refseq, query);
AlignmentUtils.VerifyNoGapsOnEnds (refseq, query);
if (callVariants) {
variants = VariantCaller.CallVariants (refseq, query, seq2.IsMarkedAsReverseComplement());
}
var newQueryQS = new QualitativeSequence (qs.Alphabet,
qs.FormatType,
query.Select (z => z.BP).ToArray (),
query.Select (p => p.QV).ToArray (),
false);
newQueryQS.Metadata = seq2.Metadata;
newQuery = newQueryQS;
} else if (seq2 is Sequence) { // For a sequence with no QV values.
var qs = seq2 as Sequence;
var query = qs.Select (v => new BPandQV (v, 0, false)).ToArray();
AlignmentUtils.LeftAlignIndels (refseq, query);
AlignmentUtils.VerifyNoGapsOnEnds (refseq, query);
// ISequence does not have a setable metadata
var newQueryS = new Sequence(qs.Alphabet, query.Select(z=>z.BP).ToArray(), false);
newQueryS.Metadata = seq2.Metadata;
if (callVariants) {
variants = VariantCaller.CallVariants (refseq, query, seq2.IsMarkedAsReverseComplement());
}
newQuery = newQueryS;
} else {
throw new ArgumentException ("Can only left align indels if the query sequence is of type Sequence or QualitativeSequence.");
}
if (aln.FirstSequence != null && aln.FirstSequence.ID != null) {
foreach (var v in variants) {
v.RefName = aln.FirstSequence.ID;
}
}
var newRef = new Sequence (seq1.Alphabet, refseq, false);
newRef.ID = seq1.ID;
newRef.Metadata = seq1.Metadata;
newQuery.ID = seq2.ID;
var newaln = new PairwiseSequenceAlignment (aln.FirstSequence, aln.SecondSequence);
var pas = new PairwiseAlignedSequence ();
pas.FirstSequence = newRef;
pas.SecondSequence = newQuery;
newaln.Add (pas);
return new Tuple<IPairwiseSequenceAlignment, List<Variant>> (newaln, variants);
}
/// <summary>
/// Convert given range containing any quality values to QualitativeSequence object
/// </summary>
/// <param name="range">Range of cells</param>
/// <param name="sequence">Sequece object</param>
/// <returns>QualitativeSequence Object</returns>
public static QualitativeSequence RangeToQualitativeSequence(List<Range> range, ISequence sequence)
{
string[] rangeData = FlattenToArray(range);
// see if we have enough quality scores to map with the sequence
if (rangeData.Length < sequence.Count)
throw new FormatException(Properties.Resources.ExportFastQ_SequenceAndScoresNotMapping);
System.Collections.IEnumerator qualityScores = rangeData.GetEnumerator();
byte[] sequenceSymbols = new byte[sequence.Count];
byte[] sequencequalityValues = new byte[sequence.Count];
byte currentQualityScore;
long curIndex = 0;
foreach (byte sequenceSymbol in sequence)
{
qualityScores.MoveNext();
if (byte.TryParse(qualityScores.Current.ToString(), out currentQualityScore))
{
sequenceSymbols[curIndex] = sequenceSymbol;
sequencequalityValues[curIndex] = currentQualityScore;
curIndex++;
}
else
{
throw new FormatException(Properties.Resources.ExportFasQ_InvalidQualityScore);
}
}
QualitativeSequence qualitativeSequence = new QualitativeSequence(sequence.Alphabet, FastQFormatType.Sanger, sequenceSymbols, sequencequalityValues)
{
ID = sequence.ID
};
return qualitativeSequence;
}
/// <summary>
/// Write quality scores to the sheet if its a FastQ file
/// </summary>
/// <param name="sequence">Sequence of which the quality scores should be written</param>
/// <param name="worksheet">Worksheet to which to write</param>
/// <param name="startingRow">Starting row</param>
/// <param name="startingColumn">Column we wrote to</param>
/// <param name="dataRange">Range where quality values were written</param>
/// <returns>Index of row after last written row</returns>
private int WriteQualityValues(
QualitativeSequence sequence,
Worksheet worksheet,
int startingRow,
int startingColumn,
out Range dataRange)
{
string[,] qualityScores = ExcelImportFormatter.FastQQualityValuesToRange(sequence, maxNumberOfCharacters);
dataRange = this.WriteToSheet(worksheet, qualityScores, startingRow, startingColumn);
return startingRow + qualityScores.GetLength(0) + 1;
}
/// <summary>
/// Return a new QualitativeSequence representing the complement of this QualitativeSequence.
/// </summary>
public ISequence GetComplementedSequence()
{
byte[] newSequenceData = new byte[this.sequenceData.GetLongLength()];
sbyte[] newQualityScores = this.qualityScores;
for (long index = 0; index < this.sequenceData.GetLongLength(); index++)
{
byte complementedSymbol;
byte symbol = this.sequenceData[index];
if (!this.Alphabet.TryGetComplementSymbol(symbol, out complementedSymbol))
{
throw new NotSupportedException(string.Format(CultureInfo.CurrentUICulture, Properties.Resource.ComplementNotSupportedByalphabet, (char)symbol, this.Alphabet.Name));
}
newSequenceData[index] = complementedSymbol;
}
QualitativeSequence seq = new QualitativeSequence(this.Alphabet, this.FormatType, newSequenceData, newQualityScores, false);
seq.ID = this.ID;
seq.metadata = this.metadata;
return seq;
}
/// <summary>
/// Return a new QualitativeSequence representing this QualitativeSequence with the orientation reversed.
/// </summary>
public ISequence GetReversedSequence()
{
byte[] newSequenceData = new byte[this.sequenceData.GetLongLength()];
sbyte[] newQualityScores = new sbyte[this.qualityScores.GetLongLength()];
for (long index = 0; index < this.sequenceData.GetLongLength(); index++)
{
newSequenceData[index] = this.sequenceData[this.sequenceData.GetLongLength() - index - 1];
newQualityScores[index] = (this.qualityScores[this.qualityScores.GetLongLength() - index - 1]);
}
QualitativeSequence seq = new QualitativeSequence(this.Alphabet, this.FormatType, newSequenceData, newQualityScores, false);
seq.ID = this.ID;
seq.metadata = this.metadata;
return seq;
}
/// <summary>
/// Converts the current instance to the specified FastQ format type
/// and returns a new instance of QualitativeSequence.
/// </summary>
/// <param name="formatType">FastQ format type to convert.</param>
public QualitativeSequence ConvertTo(FastQFormatType formatType)
{
sbyte[] convertedQualityScores = ConvertQualityScores(this.FormatType, formatType, this.qualityScores);
QualitativeSequence seq = new QualitativeSequence(this.Alphabet, formatType, this.sequenceData, convertedQualityScores, false);
seq.ID = this.ID;
seq.metadata = this.metadata;
return seq;
}
/// <summary>
/// Iterate through sequenceDictionary once only, and perform operations on each sequence list
/// Set the average read and alignment qualities for all distinct reads, and the frequency of occurrence of each
/// distinct read
/// </summary>
private void IterateSequenceDict()
{
double[] alignmentQualities = new double[CountDistinct]; // one for every sequence in the map, just get its qualities
double[] readQualities = new double[CountDistinct]; //
Collection<int> frequencies = new Collection<int>();
int i = 0;
foreach (List<SAMAlignedSequence> seqList in sequenceDict.Values)
{
// Alighment qualities
alignmentQualities[i] = seqList[0].MapQ;
// Read qualities
QualitativeSequence qSeq = new QualitativeSequence(SAMDnaAlphabet.Instance, FastQFormatType.Sanger, GetSequence(seqList[0]), GetReadQuality(seqList[0]));
readQualities[i++] = qSeq.GetQualityScores().Average();
// Frequencies
frequencies.Add(seqList.Count);
}
alignmentQuality = alignmentQualities.Length > 0 ? Math.Round(alignmentQualities.Average(), 2) : 0;
readQuality = readQualities.Length > 0 ? Math.Round(readQualities.Average(), 2) : 0;
//frequencyDistributionSequences = frequencies;
}
/* Cannot override, because base is not virtual
public override IEnumerable<QualitativeSequence> Parse(StreamReader reader)
{
FastQFormatType formatType = this.FormatType;
do
{
var seq = ParseOne(reader, formatType);
if (seq != null)
yield return ParseHeader(seq);
}
while (!reader.EndOfStream);
}*/
private QualitativeSequence ParseHeader(QualitativeSequence seq)
{
Match m = pre18Regex.Match(seq.ID);
if (m.Success)
{
seq.Metadata["Instrument"] = m.Captures[0].Value;
seq.Metadata["Lane"] = Int32.Parse(m.Captures[1].Value);
seq.Metadata["Tile"] = Int32.Parse(m.Captures[2].Value);
seq.Metadata["X"] = Int32.Parse(m.Captures[3].Value);
seq.Metadata["Y"] = Int32.Parse(m.Captures[4].Value);
seq.Metadata["Index"] = Int32.Parse(m.Captures[5].Value);
seq.Metadata["PairMember"] = Int32.Parse(m.Captures[6].Value);
return seq;
}
else
{
m = post18Regex.Match(seq.ID);
if (m.Success)
{
seq.Metadata["Instrument"] = m.Captures[0].Value;
seq.Metadata["Run"] = Int32.Parse(m.Captures[1].Value);
seq.Metadata["FlowCell"] = m.Captures[2].Value;
seq.Metadata["Lane"] = Int32.Parse(m.Captures[3].Value);
seq.Metadata["Tile"] = Int32.Parse(m.Captures[4].Value);
seq.Metadata["X"] = Int32.Parse(m.Captures[5].Value);
seq.Metadata["Y"] = Int32.Parse(m.Captures[6].Value);
seq.Metadata["PairMember"] = Int32.Parse(m.Captures[7].Value);
seq.Metadata["IsFiltered"] = m.Captures[8].Value == "Y";
seq.Metadata["ControlBits"] = Int32.Parse(m.Captures[9].Value);
seq.Metadata["IndexSequence"] = m.Captures[10].Value;
return seq;
}
}
throw new FileFormatException("Sequence identifier not in Illumina format");
}
