Alignment & variants

Variant to construct to genotyping bridge

Turn one variant into one buildable plan: verify the reference allele actually sits where the coordinate says, apply the edit, design site-directed mutagenesis primers to install it, design KASP/ARMS allele-specific primers to genotype it afterwards, and consolidate everything into a single oligo order table. Takes either a construct sequence with a 1-based position and ref/alt alleles (offline, deterministic), or an HGVS "c." description resolved through the MANE crosswalk and a live Ensembl exon map. A mismatched reference allele is refused with the bases that were actually found there, because a coordinate that is right for another isoform yields a perfectly valid primer set for the wrong base. Bases shared by both alleles are trimmed first, so a VCF-anchored pair is designed as the substitution or indel it actually is. Mutagenesis covers every class (a substitution, an insertion, a deletion and a multi-base replacement are all one interval replacement); KASP needs a single-base substitution's 3'-terminal base, so for an indel the genotyping half comes back as a named omission with the reason and the readout that does work, never as an empty list.

Not a workflow step (it does not take a single sequence argument), but its editedSequence field is the sequence to carry into the next call yourself.

post/variant_to_construct

Request

  • Base URL: https://seqbench.com/api/v1
  • URL: https://seqbench.com/api/v1/variant_to_construct
  • Auth: none required

Request body

sequencestring

OFFLINE MODE. The reference/construct sequence the variant is described against (raw or FASTA). Mutually exclusive with "variant".

positioninteger

OFFLINE MODE. 1-based position of the first base of refAllele on sequence. For a pure insertion (empty refAllele) the alternate bases are inserted immediately BEFORE this position.

refAllelestring

OFFLINE MODE. Bases replaced, as they appear on sequence — checked against it and refused if they differ. Use "-" or "" for a pure insertion. A VCF-style anchored pair (both alleles carrying a shared flanking base, e.g. refAllele "T" / altAllele "TGGATCC", or "TG" / "AG") is accepted and trimmed to its minimal form before anything is designed; the trimmed coordinates are reported as minimal.

altAllelestring

OFFLINE MODE. Bases installed. Use "-" or "" for a pure deletion.

variantstring

NETWORK MODE. A full HGVS "c." description, e.g. "NM_000546.6:c.215C>G" or "TP53:c.215C>G". Resolved through the bundled MANE crosswalk and a live Ensembl exon map, then a genomic window is fetched around it. Mutually exclusive with "sequence".

flankinteger

NETWORK MODE. Bases of genomic context fetched either side of the variant (60-1000). Needs to cover the allele-specific core upstream and the genotyping amplicon downstream.

labelstring

Name stem for the ordered oligos. Defaults to the HGVS string (network mode) or a positional label like "A100G" (offline).

style'quikchange' | 'q5'

Mutagenic primer style: overlapping (QuikChange) or back-to-back (Q5/KLD).

armTmTargetnumber

Target Tm (°C) for each mutagenic primer's template-binding arm (45-80; the arm is grown between 10 and 30 nt, so a target outside that range is refused rather than clamped to the shortest or longest arm).

targetCoreTmnumber

Target Tm (°C) for the allele-specific genotyping core, before the universal tail (45-80; the core is chosen between 18 and 27 nt, so a target outside that range is refused rather than clamped).

minAmpliconinteger

Minimum genotyping amplicon length, measured from the allele-specific primer's 5' end (which sits one core-length upstream of the variant) to the reverse primer's 5' end. Must not exceed maxAmplicon.

maxAmpliconinteger

Maximum genotyping amplicon length. Refused below 36 bp, the shortest KASP product that can exist (an 18 nt core plus an 18 nt reverse primer), and refused below minAmplicon — both are unsatisfiable for every reference, and left unchecked they came back as "no common reverse primer could be placed; provide more 3' flanking sequence".

addSecondaryMismatchboolean

Engineer the ARMS destabilizing mismatch 3 nt from the allele-specific primer's 3' end.

Response

Tool ran successfully.

okunknown required
toolstring required

The tool slug that ran.

resultobject required

Tool-specific output object. Its shape depends on the tool — call GET /{tool} or see the docs for each tool's fields.

Example response

{
  "provenance": {
    "apiVersion": "1.1.0"
  }
}

Changes

No recorded changes to this operation across all 1 revision of this API.