/// <summary>
/// Make sure the aligner is supported
/// </summary>
/// <param name="aligner"></param>
public static string GetAlignerOption(string spritzDirectory, RSEMAlignerOption aligner)
{
if (aligner == RSEMAlignerOption.Bowtie1)
{
throw new NotSupportedException("Use of Bowtie1 is not supported. Use STAR or Bowtie2 instead.");
}
string alignerOption = aligner == RSEMAlignerOption.STAR ? "--star" : // STAR is in the path, now
aligner == RSEMAlignerOption.Bowtie2 ? "--bowtie2 --bowtie2-path " + WrapperUtility.ConvertWindowsPath(TopHatWrapper.GetBowtie2DirectoryPath(spritzDirectory)) :
null;
return(alignerOption);
}
public TranscriptQuantificationParameters(string spritzDirectory, string analysisDirectory, string referenceFastaPath, int threads, string geneModelPath,
RSEMAlignerOption aligner, Strandedness strandedness, string[] fastq, bool doOutputBam)
{
SpritzDirectory = spritzDirectory;
AnalysisDirectory = analysisDirectory;
ReferenceFastaPath = referenceFastaPath;
Threads = threads;
GeneModelPath = geneModelPath;
Aligner = aligner;
Strandedness = strandedness;
Fastq = fastq;
DoOutputQuantificationBam = doOutputBam;
}
/// <summary>
/// Gets commands to calculate expression an RSEM reference
/// </summary>
/// <param name="spritzDirectory"></param>
/// <returns></returns>
public List <string> CalculateExpressionCommands(string spritzDirectory, string referencePrefix, int threads, RSEMAlignerOption aligner, Strandedness strandedness,
string[] fastqPaths, bool doOuptutBam)
{
if (fastqPaths.Length < 1)
{
throw new ArgumentOutOfRangeException("No fastq files were given for RSEM calculate expression.");
}
if (fastqPaths.Length > 2)
{
throw new ArgumentOutOfRangeException("Too many fastq file types given for RSEM calculate expression.");
}
List <string> scriptCommands = new List <string>
{
WrapperUtility.ChangeToToolsDirectoryCommand(spritzDirectory),
"cd RSEM-1.3.0",
};
string[] analysisFastqPaths = fastqPaths;
string alignerOption = GetAlignerOption(spritzDirectory, aligner);
string threadOption = "--num-threads " + threads.ToString();
string strandOption = "--strandedness " + strandedness.ToString().ToLowerInvariant();
// Decompress files if needed
// The '--star-gzipped-read-file' and '--star-bzipped-read-file' options work, but then the rest of RSEM doesn't when using compressed files...
bool fastqIsGunzipped = analysisFastqPaths[0].EndsWith("gz");
bool fastqIsBunzipped = analysisFastqPaths[0].EndsWith("bz2") || analysisFastqPaths[0].EndsWith("bz");
if (fastqIsGunzipped || fastqIsBunzipped)
{
for (int i = 0; i < analysisFastqPaths.Length; i++)
{
string decompressionCommand = fastqIsGunzipped ? "gunzip" : "bunzip2";
scriptCommands.Add($"{decompressionCommand} --keep {WrapperUtility.ConvertWindowsPath(analysisFastqPaths[i])}");
analysisFastqPaths[i] = Path.ChangeExtension(analysisFastqPaths[i], null);
}
}
string inputOption = analysisFastqPaths.Length == 1 ? string.Join(",", analysisFastqPaths[0].Split(',').Select(f => WrapperUtility.ConvertWindowsPath(f))) :
"--paired-end " +
string.Join(",", analysisFastqPaths[0].Split(',').Select(f => WrapperUtility.ConvertWindowsPath(f))) +
" " +
string.Join(",", analysisFastqPaths[1].Split(',').Select(f => WrapperUtility.ConvertWindowsPath(f)));
var megabytes = Math.Floor((double)Process.GetCurrentProcess().VirtualMemorySize64 / 1000000);
string bamOption = doOuptutBam ? "--output-genome-bam" : "--no-bam-output";
OutputPrefix = Path.Combine(Path.GetDirectoryName(analysisFastqPaths[0].Split(',')[0]),
Path.GetFileNameWithoutExtension(analysisFastqPaths[0].Split(',')[0]) +
"_" + Path.GetExtension(analysisFastqPaths[0].Split(',')[0]).Substring(1).ToUpperInvariant() +
referencePrefix.GetHashCode().ToString());
// RSEM likes to sort the transcript.bam file, which takes forever and isn't very useful, I've found. Just sort the genome.bam file instead
string samtoolsCommands = !doOuptutBam ?
"" :
"if [[ ! -f " + WrapperUtility.ConvertWindowsPath(OutputPrefix + GenomeSortedBamSuffix) + " && ! -s " + WrapperUtility.ConvertWindowsPath(OutputPrefix + GenomeSortedBamSuffix) + " ]]; then\n" +
" " + SamtoolsWrapper.SortBam(OutputPrefix + GenomeBamSuffix, threads) + "\n" +
" " + SamtoolsWrapper.IndexBamCommand(OutputPrefix + GenomeSortedBamSuffix) + "\n" +
"fi";
// construct the commands
scriptCommands.AddRange(new List <string>
{
"if [[ ! -f " + WrapperUtility.ConvertWindowsPath(OutputPrefix + IsoformResultsSuffix) + " && ! -s " + WrapperUtility.ConvertWindowsPath(OutputPrefix + IsoformResultsSuffix) + " ]]; then " +
"./rsem-calculate-expression " +
"--time " + // include timed results
"--calc-ci " + // posterior calculation of 95% confidence intervals
alignerOption + " " +
threadOption + " " +
bamOption + " " +
inputOption + " " +
WrapperUtility.ConvertWindowsPath(referencePrefix) + " " +
WrapperUtility.ConvertWindowsPath(OutputPrefix) +
"; fi",
samtoolsCommands
});
return(scriptCommands);
}
/// <summary>
/// Gets commands to prepare an RSEM reference
/// </summary>
/// <param name="spritzDirectory"></param>
/// <param name="referenceFastaPath"></param>
/// <param name="referencePrefix"></param>
/// <param name="threads"></param>
/// <param name="geneModelPath"></param>
/// <param name="aligner"></param>
/// <returns></returns>
public List <string> PrepareReferenceCommands(string spritzDirectory, string referenceFastaPath, int threads, string geneModelPath, RSEMAlignerOption aligner)
{
// make option strings, including putting reference files into a new directory
string alignerOption = GetAlignerOption(spritzDirectory, aligner);
string threadOption = "--num-threads " + threads.ToString();
string referencePrefixDirectory = Path.Combine(Path.GetDirectoryName(referenceFastaPath), Path.GetFileNameWithoutExtension(referenceFastaPath)) +
(aligner == RSEMAlignerOption.STAR ? "RsemStarReference" : "RsemBowtieReference") +
"_" + Path.GetExtension(geneModelPath).Substring(1).ToUpperInvariant() + geneModelPath.GetHashCode().ToString();
ReferenceIndexPrefix = Path.Combine(referencePrefixDirectory, Path.GetFileNameWithoutExtension(referenceFastaPath));
string geneModelOption = Path.GetExtension(geneModelPath).StartsWith(".gff") ? "--gff3 " + WrapperUtility.ConvertWindowsPath(geneModelPath) :
Path.GetExtension(geneModelPath) == ".gtf" ? "--gtf " + WrapperUtility.ConvertWindowsPath(geneModelPath) :
null;
// construct the commands
var scriptStrings = new List <string>
{
WrapperUtility.ChangeToToolsDirectoryCommand(spritzDirectory),
"cd RSEM-1.3.0",
"mkdir " + WrapperUtility.ConvertWindowsPath(referencePrefixDirectory),
"if [[ ! -f " + WrapperUtility.ConvertWindowsPath(Path.Combine(referencePrefixDirectory, "SA")) + " && ! -s " + WrapperUtility.ConvertWindowsPath(Path.Combine(referencePrefixDirectory, "SA")) + " ]]; then " +
"./rsem-prepare-reference " +
geneModelOption + " " +
alignerOption + " " +
threadOption + " " +
WrapperUtility.ConvertWindowsPath(referenceFastaPath) + " " +
WrapperUtility.ConvertWindowsPath(ReferenceIndexPrefix) +
"; fi"
};
return(scriptStrings);
}
