GotCloud: FAQs

GotCloud Frequently Asked Questions (FAQs)

This page hopes to answer some of the FAQs about GotCloud.

If you cannot find the answer to your question here, please contact us.

Compiling FAQs

How do I compile

cd src/; make;

CMake Error: The current CMakeCache.txt directory /path1/gotcloud/src/premo/build/CMakeCache.txt is different than the directory /path2/gotcloud/src/premo/build where CMakeCache.txt was created. This may result in binaries being created in the wrong place. If you are not sure, reedit the CMakeCache.txt
CMake Error: The source "/path1/gotcloud/src/premo/CMakeLists.txt" does not match the source "/path2/gotcloud/src/premo/CMakeLists.txt" used to generate cache. Re-run cmake with a different source directory.

This compilation error likely occurred because you moved GotCloud to a new directory and premo (one of the tools included in GotCloud) was still pointing to the old location.

• Remove src/premo/build/CMakeCache.txt and recompile.

Alignment Pipeline FAQs

Error "read <> mapped to 'chr1' at POS 0 to -1, flag 69,0 has BIN 4680 but should be 266824".
        "Fix it by using BAM->SAM->BAM to force a recalculation of the BIN field."
        "Fail to index the BAM file"

This error appears to be due to BWA reference files that do not match the version of BWA used to run GotCloud align. Run the following to regenerate the BWA reference files with the GotCloud version of BWA.

gotcloud/bin/bwa index path/to/your/ref.fa

Variant Calling Pipeline FAQs

Targetted/Exome

Exiting due to ERROR:
        INVALID_SORT: ERROR: File is not coordinate sorted at record 1003
        PreviousRecord was 1:15347, but this record is 1:15328

This error occurs if you have SAMTOOLS_VIEW_TARGET_ONLY = TRUE in your configuration file and a read(s) in your BAM file overlaps multiple regions in your BED file.

To fix this, set:

SAMTOOLS_VIEW_TARGET_ONLY = FALSE

Further Explanation of the issue:

The SAMTOOLS_VIEW_TARGET_ONLY set to TRUE tells our pipeline to pass to SAMTOOLS view all of the regions in the BED file that fall into a single segment of our processing.

It appears that samtools view will then go through each of those regions independently and one at a time, selecting just the records that fall within that region.

A problem occurs if records fall within multiple regions from the BED file. Since samtools view process the regions independently, it will write those records multiple times and they will end up out of order.

For example:

If we have BED with regions chr20 20300049-20300050 and chr20 20300052-20300053

The resulting call to samtools view looks like this:

gotcloud/bin/samtools-hybrid view -q 20 -F 0x0704 bamFile.bam 20:20300049-20300050 20:20300052-20300053

Say there are reads at positions 20299969 and 20299994 that are each 76 bases long, they will end up being output twice by the samtools view call and in this order:

20299969

20299994

20299969

20299994

Version Problems

Version 1.08

ERROR: Could not find umake/outputDir in currentDirectory

To work around this bug, specify an absolute path for OUT_DIR/--outdir

This is due to the aligner trying to use OUT_PREFIX to transform the output directory to an absolute path, by prepending OUT_PREFIX. But, OUT_PREFIX is a snpcall setting that sets the snpcall Makefile filename. OUT_PREFIX will be deprecated in the next release. The current OUT_PREFIX which refers to the snpcall Makefile filename will be renamed to something else.

Version 1.07

Type of arg 1 to keys must be hash (not hash element) at gotcloud/bin/Conf.pm line 155, near "}) "
Compilation failed in require at gotcloud/bin/align.pl line 56.

To fix this bug, you need to edit gotcloud/bin/Conf.pm, line 155. This error only occurs with certain versions of perl.

Change

foreach my $key (keys $CONF_HASH{$section}) {

to

foreach my $key (keys %{$CONF_HASH{$section}}) {

Illegal division by zero at gotcloud/scripts/vcf-summary line 388.
make: *** [vcfs/chrY/chrY.hardfiltered.vcf.gz.OK] Error 255
make: *** Waiting for unfinished jobs....
Commands finished in xxxxx secs WITH ERRORS. Check the logs

The current version does not gracefully handle a chromosome with no variants.

By default, GotCloud assumes all samples are female, producing no chr Y variants, which will produce this error.

Work arounds:

• Disable Chromosome Y by adding the following to your configuration file:

CHRS = 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 X

You may also want to disable chr X since it may be slightly misleading due to the assumption that all samples are female.

• Specify a pedigree file that tells GotCloud which individuals are Male & which are Female by adding the following to your configuration file:

PED_INDEX = pedigreeFile.ped

GotCloud only looks at the 2nd & 5th columns of the pedigree file (white-space delimited)

• 2nd column is the individual ID which should match the 1st column of the BAM_INDEX file.
• 5th column is the sex, use: 1 for male and 2 for female.