pipline_workflow.dot

digraph sns_pipeline {
// overall graph label
// label="NGS580 Data Analysis Pipeline\n\n"
label=<<FONT POINT-SIZE="20">NGS580 Data Analysis Pipeline</FONT><BR /><BR />>
labelloc="t";

graph [fontname = "helvetica"];
node [fontname = "helvetica"];
edge [fontname = "helvetica"];

// define all the nodes in the order of appearance
Start
Trimmomatic
BWA_MEM
Sambamba
Rsamtools
GATK_DepthOfCoverage
GATK_Realign
GATK_Recalibrate
CNVkit
Control_Freec
CNV_compare
LoFreq
GATK_HaplotypeCaller
GATK_MuTect2
ANNOVAR
Reporting
QC
Review

// add styles and labels to the pipeline steps
Start [
    label=<<FONT POINT-SIZE="14">
    Demultiplexed sequencing<BR />
    reads from NextSeq<BR />
    &#8226; output: .fastq
    </FONT>>,
    style=rounded, shape=box, color=Black
    ]

// pre-processing pipeline steps
Trimmomatic [
    label=<<FONT POINT-SIZE="18">Trimmomatic</FONT><BR />
    <FONT POINT-SIZE="12">
    trim low quality sequence reads<BR />
    &#8226; input: .fastq<BR />
    &#8226; output: .fastq
    </FONT>>,
    style=rounded, shape=box, color=Blue
    ]

BWA_MEM [
    label=<<FONT POINT-SIZE="18">BWA MEM</FONT><BR />
    <FONT POINT-SIZE="12">
    align reads<BR />
    &#8226; input: .fastq<BR />
    &#8226; output: .bam
    </FONT>>,
    style=rounded, shape=box, color=Blue
    ]

Sambamba [
    label=<<FONT POINT-SIZE="18">Sambamba</FONT><BR />
    <FONT POINT-SIZE="12">
    filter poorly mapped and duplicated reads<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .bam
    </FONT>>,
    style=rounded, shape=box, color=Blue
    ]

GATK_Realign [
    label=<<FONT POINT-SIZE="18">GATK</FONT><BR />
    <FONT POINT-SIZE="16">
    RealignerTargetCreator<BR />
    IndelRealigner<BR />
    BaseRecalibrator<BR />
    </FONT>
    <FONT POINT-SIZE="12">
    recalibrate and realign reads<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .bam
    </FONT>>,
    style=rounded, shape=box, color=Blue
    ]

GATK_Recalibrate [
    label=<<FONT POINT-SIZE="18">GATK</FONT><BR />
    <FONT POINT-SIZE="16">
    AnalyzeCovariates<BR />
    PrintReads<BR />
    BaseRecalibrator<BR />
    </FONT>
    <FONT POINT-SIZE="12">
    visualize and save recalibration results<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .bam, .txt, .pdf
    </FONT>>,
    style=rounded, shape=box, color=Blue
    ]

// quality control pipeline steps -
Rsamtools [
    label=<<FONT POINT-SIZE="18">Rsamtools</FONT><BR />
    <FONT POINT-SIZE="12">
    calculate and visualize fragment sizes<BR />
    (custom scripts)<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .pdf
    </FONT>>,
    style=rounded, shape=box, color=Orange
    ]

GATK_DepthOfCoverage [
    label=<<FONT POINT-SIZE="18">GATK<BR /></FONT>
    <FONT POINT-SIZE="16">
    DepthOfCoverage
    </FONT>
    <FONT POINT-SIZE="12">
    <BR />evaluate coverage at target regions<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .txt
    </FONT>>,
    style=rounded, shape=box, color=Orange
    ]


// Variant Calling & analysis steps
LoFreq [
    label=<<FONT POINT-SIZE="18">LoFreq</FONT><BR />
    <FONT POINT-SIZE="12">
    high sensitivity variant calling<BR />
    &#8226; input: .bam, .bed<BR />
    &#8226; output: .vcf
    </FONT>>,
    style=rounded, shape=box, color=Green
    ]

GATK_HaplotypeCaller [
    label=<<FONT POINT-SIZE="18">GATK<BR />HaplotypeCaller</FONT><BR />
    <FONT POINT-SIZE="12">
    variant calling<BR />
    &#8226; input: .bam, .bed<BR />
    &#8226; output: .vcf
    </FONT>>,
    style=rounded, shape=box, color=Green
    ]

GATK_MuTect2 [
    label=<<FONT POINT-SIZE="18">GATK<BR />MuTect2</FONT><BR />
    <FONT POINT-SIZE="12">
    tumor-normal variant calling<BR />
    &#8226; input: .bam, .bed<BR />
    &#8226; output: .vcf
    </FONT>>,
    style=rounded, shape=box, color=Green
    ]

CNVkit [
    label=<<FONT POINT-SIZE="18">CNVKit</FONT><BR />
    <FONT POINT-SIZE="12">
    copy number variant analysis<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .txt, .pdf
    </FONT>>,
    style=rounded, shape=box, color=Green
]

Control_Freec [
    label=<<FONT POINT-SIZE="18">Control-Freec</FONT><BR />
    <FONT POINT-SIZE="12">
    copy number variant analysis<BR />
    &#8226; input: .bam<BR />
    &#8226; output: .txt, .pdf
    </FONT>>,
    style=rounded, shape=box, color=Green
]

ANNOVAR [
    label=<<FONT POINT-SIZE="18">ANNOVAR</FONT><BR />
    <FONT POINT-SIZE="12">
    Variant annotation<BR />
    &#8226; input: .vcf<BR />
    &#8226; output: .txt
    </FONT>>,
    style=rounded, shape=box, color=Green
    ]

// custom downstream steps
Reporting [
    label=<<FONT POINT-SIZE="14">Custom Report Generation</FONT>>,
    style=rounded, shape=box, color=Black
    ]

QC [
    label=<<FONT POINT-SIZE="14">QC Check and email output</FONT>>,
    style=rounded, shape=box, color=Black
    ]

CNV_compare [
    label=<<FONT POINT-SIZE="14">Compare CNV analysis output</FONT>>,
    style=rounded, shape=box, color=Black
    ]

Review [
    label=<<FONT POINT-SIZE="14">Clinical Review</FONT>>,
    style=rounded, shape=box, color=Purple
    ]

// layout
subgraph cluster1 {
    style="invis"
    Trimmomatic -> BWA_MEM -> Sambamba
    Sambamba -> Rsamtools
    Sambamba -> GATK_DepthOfCoverage
    Sambamba -> CNVkit
    Sambamba -> Control_Freec

    }

subgraph cluster2 {
    style="invis"
     GATK_Realign -> GATK_Recalibrate
     GATK_Recalibrate -> LoFreq -> ANNOVAR
     GATK_Recalibrate -> GATK_HaplotypeCaller -> ANNOVAR
     GATK_Recalibrate -> GATK_MuTect2 -> ANNOVAR
    }

subgraph cluster3 {
    style="invis"
    Reporting -> CNV_compare
    Reporting -> QC
    CNV_compare -> Review
    Reporting -> Review
    QC -> Review
}

splines="ortho"

{rank=same Start Trimmomatic GATK_Realign}
Start -> Trimmomatic

// Sambamba -> CNVkit
// Sambamba -> Control_Freec

CNVkit -> CNV_compare
Control_Freec -> CNV_compare

// CNV_compare -> Reporting

Sambamba -> GATK_Realign //[constraint=false]
ANNOVAR -> Reporting

Rsamtools -> Reporting
GATK_DepthOfCoverage -> Reporting
{rank=same QC Review}

// Reporting -> QC
// Reporting -> Review
// QC -> Review

}