Counting number of NSMs from WES variants

Question

[tomer9w]

Hi all!! I'm analyzing WES, currently holding a vcf file with all variant from a particular sample. I would like to annotate and count number of non-synonymous mutations to compare between the samples. Few question for you guys:)

1. I saw that SNPEFF is a good tool for annotation, but how can i use it in order to count NSMs?
2. Wanted to verify my analysis so far, just to make sure it makes sense:

alignment using BWA bwa mem -t 20 ref/gencode.vM25.transcripts.fa.gz Raw/Mel_B16-F10_1.fastq Raw/Mel_B16-F10_2.fastq > BWA/Mel_B16-F10.sam

converting to BAM, sorting and indexing: samtools view -@ 10 -b BWA/Mel_B16-F10.sam > BWA/Mel_B16-F10.bam samtools sort BWA/Mel_B16-F10.bam -@ 10 -o BWA/Mel_B16-F10.sorted.bam samtools index BWA/Mel_B16-F10.sorted.bam -@ 10

variant calling: bcftools mpileup --threads 10 -Ov -f ref/gencode.vM27.transcripts.fa BWA/Mel_B16-F10.sorted.bam | bcftools call --threads 10 -mvO z -o VCF/Mel_B16-F10.vcf

Does this pipeline makes sense?

1. How can i filter out variant with Phred-scaled quality score <30 ?

Thank you all!