[nilshomer]

Nils, congratulations on getting the publication out!

I'm about to give this a try on an odd data set -- 2kb of genomic sequence at an average (but far from uniform) coverage of around 100.000 X. It's a sequencing mixture, and the lower cutoff of variation we'd like to be able to detect is at around 0.5% (after error correction) or 500 observations.

Other than the biological samples we also have a mix of known genomic frequencies and defined indel regions to optimize parameters. Can you think of a realistic set of starting parameters?

It wasn't designed for such high coverage so all bets are off.

[apratap]

Hi Nils

Just wondering can SRMA be used for rescuing orphaned reads. So we have a dataset of variable insert library as we are sequencing the 5' and 3' end of transcripts. As a result the distance between the mates( <--- --->) is dependent on the length of transcript. To map the reads initially I am first using Mosaik which i belv does a better job with variable insert mate pair data.

After mapping we still see 40% orphaned reads where one read maps and the other doesn't. I am wondering if SRMA can rescue these reads.

Thanks!
-Abhi

[nilshomer]

No, SRMA is not for read rescue. It is for re-aligning the reads to create a better consensus.

[apratap]

Ok good to know. I will start a new thread for my question then.

Best,
-Abhi

[ymc]

Dead project now? Are there other alternatives that work on the whole genome?

[nilshomer]

Dead project now? Are there other alternatives that work on the whole genome?

It's not a dead project, feel free to post questions and bug reports etc.

[adaptivegenome]

Dead project now? Are there other alternatives that work on the whole genome?

I have used it and it is fast. I have sometimes had trouble with files in the 100GB range but generally it works fine.

We have also parallelized the GATK implementation of LR if you are interested. I am not sure which is better at realigning. I do remember comparing SRMA and GATK LR and there are differences but it was not clear to me if one was consistently better than the other. I suspect that Nils would be a better source for info on that.

[ymc]

Tried several bams with 0.1.16 but all I got was this:

at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)

[nilshomer]

Tried several bams with 0.1.16 but all I got was this:

at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)
at java.util.ArrayList$SubList.add(ArrayList.java:965)

Could you post the full error message?

[colindaven]

I have been interested in this tool for some time but never got it working:
Input is a sorted bam.

java -Xmx16g -jar srma-0.1.15.jar I=491_full_s.bam O=srma_491.bam R=../NC_002516.fna
[Fri Aug 17 10:00:54 CEST 2012] srma.SRMA INPUT=[491_full_s.bam] OUTPUT=[srma_491.bam] REFERENCE=../NC_002516.fna OFFSET=20 MIN_MAPQ=0 MINIMUM_ALLELE_PROBABILITY=0.1 MINIMUM_ALLELE_COVERAGE=3 MAXIMUM_TOTAL_COVERAGE=100 CORRECT_BASES=false USE_SEQUENCE_QUALITIES=true QUIET_STDERR=false MAX_HEAP_SIZE=8192 MAX_QUEUE_SIZE=65536 GRAPH_PRUNING=false NUM_THREADS=1 TMP_DIR=/tmp/colin2 VERBOSITY=INFO QUIET=false VALIDATION_STRINGENCY=STRICT COMPRESSION_LEVEL=5 MAX_RECORDS_IN_RAM=500000 CREATE_INDEX=false CREATE_MD5_FILE=false
java.util.NoSuchElementException
at java.util.Scanner.nextLine(Scanner.java:1503)
at net.sf.picard.reference.FastaSequenceIndex.parseIndexFile(FastaSequenceIndex.java:131)
at net.sf.picard.reference.FastaSequenceIndex.<init>(FastaSequenceIndex.java:55)
at net.sf.picard.reference.IndexedFastaSequenceFile.<init>(IndexedFastaSequenceFile.java:95)
at srma.SRMA.doWork(SRMA.java:131)
at net.sf.picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:156)
at srma.SRMA.main(SRMA.java:98)
Please report bugs to [email protected]


The fasta index file looks like this:

more ../NC_002516.fna.fai
NC_002516.2 6264404 58 70 71

Cheers for any help.

[ymc]

There are thousands of lines of these error messages. If I copy the stderr output, it will be too many lines. You can replicate my problem by downloading the pair-ended reads from

ftp://ftp.1000genomes.ebi.ac.uk/vol1...sequence_read/

and then align them using bwa. I got the same bug with SRR098401_*.filt.fastq.gz and SRR035330_*.filt.fastq.gz

[nilshomer]

I have been interested in this tool for some time but never got it working:
Input is a sorted bam.

java -Xmx16g -jar srma-0.1.15.jar I=491_full_s.bam O=srma_491.bam R=../NC_002516.fna
[Fri Aug 17 10:00:54 CEST 2012] srma.SRMA INPUT=[491_full_s.bam] OUTPUT=[srma_491.bam] REFERENCE=../NC_002516.fna OFFSET=20 MIN_MAPQ=0 MINIMUM_ALLELE_PROBABILITY=0.1 MINIMUM_ALLELE_COVERAGE=3 MAXIMUM_TOTAL_COVERAGE=100 CORRECT_BASES=false USE_SEQUENCE_QUALITIES=true QUIET_STDERR=false MAX_HEAP_SIZE=8192 MAX_QUEUE_SIZE=65536 GRAPH_PRUNING=false NUM_THREADS=1 TMP_DIR=/tmp/colin2 VERBOSITY=INFO QUIET=false VALIDATION_STRINGENCY=STRICT COMPRESSION_LEVEL=5 MAX_RECORDS_IN_RAM=500000 CREATE_INDEX=false CREATE_MD5_FILE=false
java.util.NoSuchElementException
at java.util.Scanner.nextLine(Scanner.java:1503)
at net.sf.picard.reference.FastaSequenceIndex.parseIndexFile(FastaSequenceIndex.java:131)
at net.sf.picard.reference.FastaSequenceIndex.<init>(FastaSequenceIndex.java:55)
at net.sf.picard.reference.IndexedFastaSequenceFile.<init>(IndexedFastaSequenceFile.java:95)
at srma.SRMA.doWork(SRMA.java:131)
at net.sf.picard.cmdline.CommandLineProgram.instanceMain(CommandLineProgram.java:156)
at srma.SRMA.main(SRMA.java:98)
Please report bugs to [email protected]


The fasta index file looks like this:

more ../NC_002516.fna.fai
NC_002516.2 6264404 58 70 71

Cheers for any help.

It looks like your FASTA index is broken. Can you try rebuilding?

There are thousands of lines of these error messages. If I copy the stderr output, it will be too many lines. You can replicate my problem by downloading the pair-ended reads from

ftp://ftp.1000genomes.ebi.ac.uk/vol1...sequence_read/

and then align them using bwa. I got the same bug with SRR098401_*.filt.fastq.gz and SRR035330_*.filt.fastq.gz

I am sorry, please try reducing your read set or the like to a manageable test case. Otherwise, I charge $5KUSD/hour

[madonjoe]

Empty VCF file

I tried to use SRMA to realign my reads and did a variant calling. However, after SRMA, which ran fine, I got an empty vcf file. Anything I can do to fix this problem?