Variant Finding parameters

You can adjust the following Variant Finding parameters to optimize your analysis results when you create or edit analysis workflows.

IMPORTANT! Use the default parameter settings unless you are an advanced user.

Parameter Name

Description

Main tab

Analysis

Data Quality Stringency

Filter: Phred-scaled minimum average evidence per read or no-call. Related VCF field: MLLD.

Allowed values: 0 to unlimited

Suggested trial value: 6.5

Downsample to Coverage

Reduce coverage in over-sampled locations to this value.

Allowed values: 1 to unlimited

Suggested trial value: 4000

SNP Min Cov Each Strand

Filter: Minimum coverage required on each strand.

Allowed values: 0 to unlimited

Suggested trial value: >=3

SNP Min Variant Score

Filter: Phred-scaled evidence that the reads support a variant above minimum frequency.

Allowed values: 0 to unlimited

Suggested trial value: 10

SNP Min Allele Freq

Frequency cutoff for supporting a variant.

Allowed values: 0 to 1.0 (decimals)

Suggested trial value: 0.0005 to 0.005 (TagSeq), 0.01 to 0.2 (other)

SNP Min Coverage

Total coverage required of reads or no-call.

Allowed values: 0 to unlimited

Suggested trial value: 2 (TagSeq), 5 to 20 (other)

SNP Strand Bias

Filter: proportion of variant alleles comes overwhelmingly from one strand. Related VCF field: STB.

Allowed values: 0.5 to 1.0 (decimals)

Suggested trial value: 0.95

INDEL Min Cov Each Strand

Filter: Minimum coverage required on each strand.

Allowed values: 0 to unlimited

Suggested trial value: >=3

INDEL Min Variant Score

Filter: Phred-scaled evidence that the reads support a variant above minimum frequency.

Allowed values: 0 to unlimited (integers)

Suggested trial value: >=10

INDEL Min Allele Freq

Frequency cutoff for supporting a variant.

Allowed values: 0 to 1.0 (decimals)

Suggested trial value: 0.0005 to 0.005 (TagSeq), 0.05 to 0.2 (other)

INDEL Min Coverage

Total coverage required of reads or no-call.

Allowed values: 0 to unlimited (integers)

Suggested trial value: 2 to 10,000 (TagSeq), 15 to 30 (other)

INDEL Strand Bias

Filter: proportion of variant alleles comes overwhelmingly from one strand. Related VCF field: STB.

Allowed values: 0.5 to 1.0 (decimals)

Suggested trial value: 0.95

Hotspot Min Cov Each Strand

Filter: Minimum coverage required on each strand.

Allowed values: 0 to unlimited (integers

Suggested trial value: 3

Hotspot Min Variant Score

Phred-scaled evidence that the reads support a variant above minimum frequency.

Allowed values: 0 to unlimited (integers)

Suggested trial value: ≥10

Hotspot Min Allele Freq

Frequency cutoff for supporting a variant.

Allowed values: 0 to 1.0 (decimals)

Suggested trial value: 0.0005 to 0.005 (TagSeq); 0.01 to 0.2 (other)

Hotspot Min Coverage

Total coverage required of reads or no-call.

Allowed values: 0 to unlimited (integers)

Suggested trial value: 2 to 10,000 (TagSeq); 5 to 20 (other)

Hotspot Strand Bias

Filter: proportion of variant alleles comes overwhelmingly from one strand. Related VCF field: STB.

Allowed values: 0.5 to 1.0 (decimals)

Suggested trial value: 0.95

PPA

Reports Possible Polyploidy Alleles (PPA) in the INFO FIELD of the VCF file. Related VCF field: PPA. Allowed values: 1 = report PPA, 0 = do not report. 1 is recommended for somatic-only and experimental research.

If PPA is set to True and Variant View advanced parameter is set to Allele View, a PPA column is included in the Analysis Results screen. A value of Yes indicates variants that are PPA allelles. No indicates variants that are not PPA alleles.

For more information, see Locus View versus Allele View of variants.

Allowed values: True or False

Prediction Precision

Number of pseudo-data-points suggesting our predictions match the measurements without bias.

Allowed values: 0 to unlimited (decimals)

Suggested trial value: 1.0

Outlier Probability

Prior probability that a read comes from some other distribution.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0.005 to 0.01

Heavy Tailed

How heavy the t-distribution tails are to allow for unusual spread in the data.

Allowed values: 0 to unlimited (decimals)

Suggested trial value: 3

Filter Unusual Predictions

Filter: predictions are distorted to fit the data more than this distance (relative to the size of the variant). Related VCF fields: FWDB, REVB [RBI = sqrt(FWDB ^ 2 + REVB ^ 2)].

Allowed values: 0 to unlimited (decimals)

Suggested trial value: 0.3 (30% of variant change size)

Filter Insertion Predictions

Filter: observed clusters deviate from predictions more than this amount (relative to the size of the variant). Related VCF fields: VARB, REFB.

Allowed values: 0 to unlimited (decimals)

Suggested trial value: 0.2 (20% of variant change size)

Filter Deletion Predictions

Filter: observed clusters deviate from predictions more than this amount (relative to the size of the variant). Related VCF fields: VARB, REFB.

Allowed values: 0 to unlimited (decimals)

Suggested trial value: 0.2 (20% of variant change size)

HP Max Length

Filter: homopolymer length involved in an in/del. Related VCF field: HRUN.

Allowed values: 1 to unlimited (integers)

Suggested trial value: 8

Do SNP Realignment

Realign reads in the vicinity of SNP candidates. Impact: True = do not realign, False = realign.

Allowed values: True or False

Suggested trial value: PGM: (germline) True, (somatic) False; Proton: (germline) False, (somatic) False.

Suppress Recalibration

Recalibration values from pipeline used or not (experimental). No related fields, changes basecalling behavior. Allowed values: True = allow recalibration, False = don't allow recalibration.

Allowed values: True or False

Suggested trial value: False

SSE Probability Threshold

Filter out variants in motifs with error rates above this.

Allowed values: 0 to 1

Suggested trial value: 0.2

MNP Min Cov Each Strand

Filter: Minimum coverage required on each strand.

Allowed values: 0 to unlimited (integers)

Suggested trial value: >=3

MNP Min Variant Score

Filter out MNPs with a QUAL score less than or equal to this Phred-scaled value.

Allowed values: 0 to unlimited

Suggested trial value: 10

MNP Min Allele Freq

Frequency cutoff for supporting a variant.

Allowed values: 0 to 1

Suggested trial value: 0.0005 to 0.005 (TagSeq); 0.01 to 0.2 (other)

MNP Min Coverage

Total coverage required of reads or no-call.

Allowed values: 0 to unlimited (integers)

Suggested trial value: 2 to 10,000 (TagSeq); 5 to 20 (other)

MNP Strand Bias

Filter: proportion of variant alleles comes overwhelmingly from one strand. Related VCF field: STB.

Allowed values: 0.5 to 1

Suggested trial value: 0.95

MNP Strand Bias Pval

Filter out mnps with pval below this [1.0] given strand bias > mnp-strand-bias.

Allowed values: 0 to 1

Suggested trial value: 1

SNP Strand Bias Pval

Filter out snps with pval below this [1.0] given strand bias > snp-strand-bias.

Allowed values: 0 to 1

Suggested trial value: 1

INDEL Strand Bias Pval

Filter out INDELs with pval below this [1.0] given strand bias > INDEL-strand-bias.

Allowed values: 0 to 1

Suggested trial value: 1

Hotspot Strand Bias Pval

Filter out hotspot variants with pval below this [1.0] given strand bias > hotspot-strand-bias.

Allowed values: 0 to 1

Suggested trial value: 1

Position Bias Reference Fraction

Skip position bias filter if (reference read count)/(reference + alt allele read count) less than or equal to this.

Allowed values: 0 to 1

Suggested trial value: 0.05

Position Bias

Filter out variants with position bias relative to soft clip ends in reads > position-bias.

Allowed values: 0 to 1

Suggested trial value: 0.75

Position Bias Pvalue

Filter out if position bias is above the Position Bias given pval less than Position Bias Pval.

Allowed values: 0 to 1

Suggested trial value: 0.05

Use position bias

Enable the position bias filter.

Allowed values: True or False

Suggested trial value: False

INDEL As HPINDEL

Apply INDEL filters to non HP INDELs.

Allowed values: True or False

Suggested trial value: False

Do MNP Realignment

Realign reads in the vicinity of candidate mnp variants.

Allowed values: True or False

Suggested trial value: True

Realignment Threshold

Maximum allowed fraction of reads where realignment causes an alignment change.

Allowed values: 0 to 1

Suggested trial value: 1

FD Nonsnp Min Var Cov

Override min_var_coverage of the flow-disrupted variants that are not SNPs (0 to disable the override). Impact: Decreasing values make variant calls less specific but more sensitive.

Allowed values: 0 to 10

Suggested trial value: 1

Read Mismatch Limit

Do not use reads with number of mismatches (where 1 gap open counts 1) above this value.

Allowed values: 0 to unlimited

Suggested trial value: 5 (TagSeq), 0 (other)

Min Cov Fraction

Do not count reads with fraction of covering any amplicons below this threshold.

Allowed values: 0 to 1

Suggested trial value: 0.9 (TagSeq), 0 (other)

Use Input Allele Only

Only consider provided alleles in the hotspots file. 0 = generate de novo candidates, 1 = hotspots only.

Allowed values: 0 to 1

Suggested trial value: 0

Liquid Biopsy and Ion AmpliSeq™ HD

ampliseq-hd

Sets defaults for counting reads in liquid biopsy runs with ampliseq-hd using "tvc consensus".

Allowed values: 0 to 1

Suggested trial value: 0

Min Family Size

Minimum number of reads with same Unique Molecular Tag required to form a functional family. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 0 to unlimited

Suggested trial value: between 3 and 7

min-mapping-qv

Minimum mapping quality value required for a read to be counted.

Allowed values: 0 to unlimited (integer)

Suggested trial value: 20 (TagSeq); 0 (other)

Poisson

Use Poisson parameter estimation to estimate count of functional families.

Allowed values: 0 to 1

Suggested trial value: 0

SNP Min Var Coverage

Minimum number of variant supporting functional families required to make a SNP call. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 2 to 10

Suggested trial value: 2

MNP Min Var Coverage

Minimum number of variant supporting functional families required to make a MNP call. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 2 to 10

Suggested trial value: 2

INDEL Min Var Coverage

Minimum number of variant supporting functional families required to make a INDEL call. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 2 to 10

Suggested trial value: 2

Hotspot Min Var Coverage

Minimum number of variant supporting functional families required to make a hotspot call. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 2 to 10

Suggested trial value: 2

INDEL Func Size Offset

Require family of size >= (min_tag_fam_size + this value) to be functional for calling HP-INDEL. Impact: Increasing values make variant calls less sensitive but more specific.

Allowed values: 0 to 4

Suggested trial value: 2

Tag Sim Max Cov

Check the similarity of UID of variant families if the variant molecular coverage is less than or equal to this value. Related VCF field: TGSM.

Allowed values: 0 to unlimited

Suggested trial value: 20

Minimum Family Coverage per Strand

Minimum required coverage of reads on each strand in a bi-directional molecular tag family.

Allowed values: 0 to unlimited

Suggested trial value: 0

Minimum Callable Probability

Minimum callable probability for LOD calculation.

Allowed values: 0 to 1

Suggested trial value: 0.98

Suppress called allele LOD

Suppress the LOD reporting of a variant allele that is called.

Allowed values: True or False

Suggested trial value: False

Advanced tab

Analysis

Allow INDELs

Enable INDELs in FreeBayes hypothesis generator. Allowed values: True = generate INDEL hypotheses, False = don't generate.

Allowed values: True or False

Suggested trial value: True

Allow SNPs

Enable SNPs in FreeBayes hypothesis generator. Allowed values: True = generate SNP hypotheses, False = don't generate.

Allowed values: True or False

Suggested trial value: True

Allow MNPs

Enable MNPs in FreeBayes hypothesis generator. Allowed values: True = generate MNP hypotheses, False = don't generate.

Allowed values: True or False

Suggested trial value: True

Allow Complex

Enable complex variants in FreeBayes hypothesis generator. Allowed values: True = generate MNP hypotheses, False = don't generate.

Allowed values: True or False

Suggested trial value: False

Minimum mapping qv

Minimum mapping QV value required for reads to be allowed into the pileup (both FreeBayes and evaluator).

Allowed values: 0 to unlimited (integer)

Suggested trial value: 4

Read SNP Limit

Read mismatch limit on number of mismatches: filter potential mis-mapped reads.

Allowed values: 0 to unlimited (integer)

Suggested trial value: 10

Read Max Mismatch Fraction

Read maximum mismatch fraction of mismatches in length of read: filter potential mis-mapped reads.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 1

Generate Min Alt Allele Freq

Generate variants with at least this frequency in the pileup.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0.02 to 0.15

Generate Min INDEL Alt Allele Freq

Generate INDEL variants with at least this frequency in the pileup.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0.02 to 0.15

Generate Min Coverage

Generate variants in locations with at least this depth of coverage.

Allowed values: 0 to unlimited

Suggested trial value: 6

Kmer Len

Size of the smallest k-mer used in assembly. Impact: Increasing values make INDEL calls less sensitive but more specific.

Allowed values: 5 to unlimited (integers)

Suggested trial value: 11 to 30

Min Var Count

Minimum support for a variant to be evaluated. Impact: Increasing values make INDEL calls less sensitive but more specific.

Allowed values: 1 to unlimited (integers)

Suggested trial value: 3 to 30

Short Suffix Match

Minimum assembled sequence match on both sides of the variant. Impact: Increasing values make INDEL calls less sensitive but more specific.

Allowed values: 2 to unlimited (integers)

Suggested trial value: between 4 and the value given to the kmer_len parameter

Min INDEL Size

Minimum size INDEL reported by assembly. Impact: Increasing values make INDEL calls less sensitive but more specific.

Allowed values: 1 to unlimited (integers)

Suggested trial value: 2 to 30

Max HP Length

Variants containing HP larger than this are not reported. Impact: Increasing values make INDEL calls more sensitive but less specific.

Allowed values: 1 to unlimited (integers)

Suggested trial value: 2 to 11

Min Var Freq

Minimum frequency of the variant to be reported. Impact: Increasing values make INDEL calls less sensitive but more specific.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0.1 to 0.4

Relative Strand Bias

Variants with strand bias above this are not reported. Impact: Increasing values make INDEL calls more sensitive but less specific.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0.6 to 1.0

Output MNV

Enables reporting of complex variants. Impact: 1 = report complex variants, 0 = don't report.

Allowed values: 0 to 1 (decimals)

Suggested trial value: 0

Variant View

Represent variants with Locus view or Allele view.

For more information, see Locus View versus Allele View of variants.

Allowed values: Locus view or Allele view

Suggested trial value: your preference

Analysis SSE File

Analysis strand specific error file. Select a mask from the list or upload a mask from Upload.