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  • Varscan-output interpretation

    Hi all,

    I am new to NGS analysis. I have used bowtie (ver:bowtie-0.12.7) for aligning reference sequence (fastq format) with two paired end files of illumina reads (fastq format).
    I used SAM tools (ver:samtools-0.1.18) and made a 'mpileup' file.
    Then I have used Varscan (ver:.v2.2.11) for variant calling. I used "mpileup2snp" command (with default parameters) to determine SNV and for heterozygosity & homozygosity.
    My basic idea is to call heterozygous SNV with conservative filters, such as coverage, variant allele
    frequency, strand representation and p-value to isolate high-confidence calls.

    Using the command "mpileup2snp", I got the output as follows. Given 1 column as an example
    Chrom - gi|371443199|gb|JH556661.1|
    Position - 2594265
    Ref- G
    Var- C
    Cons:Cov:Reads1:Reads2:Freq:P-value- S:21:14:7:33.33%:4.3101E-3
    StrandFilter:R1+:R1-:R2+:R2-val- Fail:12:2:0:7:3.096E-4
    SamplesRef- 0
    SamplesHet- 1
    SamplesHom- 0
    SamplesNC - 0
    Cons:Cov:Reads1:Reads2:Freq:P-value- S:21:14:7:33.33%:4.3101E-3

    1.Please let me know whether I am using the correct command to call heterzygous SNV with high-confidence calls(whether I have to use mpileup2snp or mpileup2indel or mpileup2cns?)

    2.Can I parse these variant call by chromosome?. How Can I do that?

    3.Is it possible to view regions of high and low heterozygosity in the genome?. If possible, how can I do that?

    Thanks in advance

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