---
title: "twinPE Studio (dual pegRNA)"
method: POST
path: "/prime_editing_twin_design"
tags: ["CRISPR"]
---

# twinPE Studio (dual pegRNA)

`POST /prime_editing_twin_design`

## Request

- Base URL: `https://seqbench.com/api/v1`
- URL: `https://seqbench.com/api/v1/prime_editing_twin_design`
- Auth: none required

Design a twinPE pegRNA pair (Anzalone et al. 2022) for a replacement too large for a single pegRNA's RTT: a left pegRNA nicks the + strand at/before the replacement window and a right pegRNA nicks the - strand at/after it, each synthesizing a new 3' flap; both flaps are truncated at a shared overlap in the middle of the new sequence so they anneal and resolve the edit without an HDR donor. Coordinates: both pegRNAs' protospacerStart/protospacerEnd/nickPosition are 1-based inclusive in the submitted PRE-EDIT target's frame (the PAM search runs on the unedited sequence, on both sides), while replaceSpan is the span of the new content in the returned editedSequence. Off-target activity is not evaluated (no in-browser reference genome).

## Request body

- object
  - `target` string, required: Forward-strand target DNA (raw or FASTA), with flanking sequence on both sides of the replacement window.
  - `replaceStart` integer, required: 1-based inclusive start of the region being replaced/deleted.
  - `replaceEnd` integer, required: 1-based inclusive end of the region being replaced/deleted. For a pure insertion, set replaceEnd = replaceStart - 1.
  - `newSequence` string, required: New sequence (forward strand) to install in place of [replaceStart, replaceEnd]. Needs >= 2 bp.
  - `overlapLength` integer: Length (bp) of the shared overlap built into both pegRNAs' 3' flaps where they meet and anneal.
  - `pbsLength` integer: Optional preferred PBS length to highlight; a full 8-17 nt sweep is always returned.

## Response `200`

Tool ran successfully.

- SuccessResponse: Standard success envelope returned by every single-tool call.
  - `ok` unknown, required
  - `tool` string, required: The tool slug that ran.
  - `result` object, required: Tool-specific output object. Its shape depends on the tool — call `GET /{tool}` or see the docs for each tool's fields.
  - `gate` GateResult
    - `pass` boolean, required: True if and only if every hard check passed.
    - `checks` GateCheck[], required
      - `id` string, required
      - `label` string, required
      - `severity` 'hard' | 'soft', required
      - `pass` boolean, required
      - `message` string, required
      - `value` number
      - `threshold` string
    - `notChecked` string[], required: Honest list of what this gate does NOT verify.
  - `provenance` Provenance, required
    - `apiVersion` string, required
    - `tool` union, required: Tool slug, or an array of slugs (one per step) for a workflow.
      - string
      - string[]
    - `generatedAt` string, date-time, required: ISO 8601 timestamp of when the result was generated.

## Other responses

- `400`: Invalid argument or the tool could not run.
- `404`: Unknown tool.
- `429`: Rate limited (metered tools only). Body carries code `rate_limited` and retryable `true`; honor the Retry-After header.
- `503`: A metered tool is momentarily at capacity. The identical request should succeed after Retry-After; body carries code `upstream_unavailable`.

---

[API](https://skmtc.dev/seqbench/apis/seqbench-api.md) · [All operations](https://skmtc.dev/seqbench/apis/seqbench-api/llms.txt) · [OpenAPI document](https://skmtc.dev/seqbench/apis/seqbench-api/revisions/132575216865?raw)