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How to generate BAM file (from fastq files (Paired-end)) compatible with CollectInsertSizeMetrics
I have two fastq files (first and second reads in paired end data in separate .fastq files). I want to convert them to BAM file so that I can use the Picard Tool CollectInsertSizeMetrics. However, I am unable to use the tool as I get the following errors
#[Mon Oct 02 18:12:59 GMT 2017] CollectInsertSizeMetrics HISTOGRAM_FILE=insert_size_histogram.pdf INPUT=input.bam OUTPUT=output_insert_size_metrics.txt DEVIATIONS=10.0 MINIMUM_PCT=0.05 METRIC_ACCUMULATION_LEVEL=[ALL_READS] INCLUDE_DUPLICATES=false ASSUME_SORTED=true STOP_AFTER=0 VERBOSITY=INFO QUIET=false VALIDATION_STRINGENCY=STRICT COMPRESSION_LEVEL=5 MAX_RECORDS_IN_RAM=500000 CREATE_INDEX=false CREATE_MD5_FILE=false GA4GH_CLIENT_SECRETS=client_secrets.json USE_JDK_DEFLATER=false USE_JDK_INFLATER=false WARNING 2017-10-02 18:12:59 SinglePassSamProgram File reports sort order 'queryname', assuming it's coordinate sorted anyway. WARNING 2017-10-02 18:12:59 CollectInsertSizeMetrics All data categories were discarded because they contained < 0.05 of the total aligned paired data. WARNING 2017-10-02 18:12:59 CollectInsertSizeMetrics Total mapped pairs in all categories: 0.0 [Mon Oct 02 18:12:59 GMT 2017] picard.analysis.CollectInsertSizeMetrics done. Elapsed time: 0.00 minutes. Runtime.totalMemory()=126877696
I can't quite understand whether the BAM file I generated using FastqToSam Tool is incompatible?
If so how should I correct it?
What SORT_ORDER should I use?
Why does " CollectInsertSizeMetrics All data categories were discarded because they contained < 0.05 of the total aligned paired data." ?
What is meant by "CollectInsertSizeMetrics Total mapped pairs in all categories: 0.0"?
Additionally, what are the attributes of the BAM file that will allow the Picard Tool CollectInsertSizeMetrics to be compatible with it? Do I need to change the SORT_ORDER when creating the BAM using FastToSam from the two fastq files (1st and 2nd reads in paired-end data)? Which of the two "queryname" or "coordinate" is correct for the task I want to perform?