Primers & oligos

Annealing temperature by polymerase

Annealing temperature and a full cycling program for a primer PAIR, under the rule the polymerase's own vendor publishes — which is not one rule, and is only defined against the vendor's own Tm. For NEB products the Tm is computed exactly as NEB's Tm calculator computes it (SantaLucia 1998 nearest-neighbor with NEB's salt correction, at that product's own buffer and primer concentration — the same primer's Tm moves ~10 °C between NEB buffers) and NEB's calculator rule is applied to it: Q5 at Tm + 1, Phusion at 0.93 × Tm + 7.5 (both ABOVE the Tm), Taq/OneTaq/LongAmp at Tm − 5, Vent/Deep Vent at Tm − 2 above 20 nt, with NEB's ceilings. Toyobo KOD One is Tm − 5 at the salt and oligo concentration its manual names, Takara PrimeSTAR Max a FIXED 55 °C the primer Tm plays no part in (its own 2(A+T)+4(G+C)−5 sets only the annealing time), and a generic Taq/Pfu row applies the Tm − 3 to − 5 rule of thumb to a default-condition nearest-neighbor Tm. Returns denaturation/annealing/extension steps, cycle count, magnesium guidance, the Tm basis and vendor document behind each figure, and the same primers under every other polymerase in the table for comparison.

post/annealing_temperature

Request

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

Request body

forwardPrimerstring required

Forward primer, 5'→3'.

reversePrimerstring required

Reverse primer, 5'→3'.

polymerasestring required

Polymerase or master mix. One of: q5, phusion, neb_taq, onetaq, vent, deepvent, longamp, primestar_max, kod_one, taq, pfu. Common product spellings are accepted (e.g. "Q5 Hot Start", "KOD One", "PrimeSTAR Max").

ampliconBpinteger

Expected product length (bp), used for the extension time. Omit to get the per-kb rate without a time.

highComplexityTemplateboolean

Genomic or otherwise complex template. Where the vendor publishes a second, slower extension rate for one, it is used.

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.