Hi, I'm currently doing a WES pipeline to identify variants in human sequences, currently using (in order of use):

1. Read QC and trimming: fastq

2. Alignment: bwa index, bwa mem, samtools view, samtools sort, and samtools index.

3. Remove PCR duplicates: picard markduplicates?

When it comes to removing PCR duplicates, I have seen that picard's markduplicate works to identify any duplicates.

java -jar MarkDuplicates.jar I=PE_samtoolssorted.bam
O=markedduplicates.bam M=markedduplicatesmetrics.txt

However when it comes to removing the PCR duplicates that are found online that just adding REMOVE_DUPLICATES=true removes them?

java -jar MarkDuplicates.jar I=PE_samtoolssorted.bam O=removedduplicates.bam M=markedduplicatesmetrics.txt
REMOVE_DUPLICATES=true 

The output of this will be a sorted bam file with the removed PCR duplicates?

Would the input for a variant caller like deepvariant, which requires a sorted bam file be this removedduplicates.bam file?

and if so, would it be this removedduplicatessorted.bam file that needs indexing for input into deepvariant rather than the original PE_samtoolssorted.bam?

Thanks! Sorry if confusing. Amy