Using CINDEL | DeepGE

Using CINDELNote: DeepCpf1 provides an accurate deep-learning based prediction of Cas12a/Cpf1 guide activity. You might want to use DeepCpf1.Load Example

Use Load Example to populate mode-specific sample inputs, then click Submit Job.Analysis Mode

Guide Finder in Long Sequence: Input a long sequence; CINDEL scans for valid TTTV windows and scores each candidate.

Single Sequence Scoring: Enter a sequence of at least 27bp, press ENTER to find 27bp windows with a TTTV PAM on either strand, and select one candidate.

Batch Scoring: Enter one 27bp sequence per line. Sequences must be 27bp with TTTV PAM.

Input MethodsGuide Finder:

Batch Scoring:

Guide Finder carries an Input Format dropdown of nine formats — FASTA, Gene Symbol, NMID, Ensembl ID, HGNC ID, Genomic Position, Edit Sequence (an unedited/edited pair), ClinVar ID and COSMIC ID — each of which resolves to the sequences the finder scans for TTTV windows. The dropdown belongs to this mode alone; Single Sequence and Batch Scoring take the 27bp candidates directly. Edit Sequence, ClinVar ID and COSMIC ID are the three formats that specify an edit, so those three also produce the homology-directed repair (HDR) donor columns described below.

Guide Finder (in FASTA format) and Batch Scoring accept either pasted text or an uploaded file. When uploading files, press the [+] button to add files. Uploaded files can be cleared by pressing either of the checkmark buttons.

Single Sequence accepts pasted text only (no file upload), where the 27bp candidate from the dropdown is selected after pressing ENTER.

SpeciesA gene identifier is looked up in Human (GENCODE v50 on GRCh38) or Mouse (GENCODE M39 on GRCm39). The selector appears with the four gene formats, and it is offered wherever the genome-wide off-target search offers Mouse, so a run that screens against the mouse genome can design in mouse as well.A mouse symbol from an older nomenclature resolves to the current one and the job log states the substitution — Olfr1 designs against Or1e16. Whichever of the four gene formats is chosen, Mouse replaces the identifier box with Enter Mouse Gene Name or ID, which takes a mouse symbol or a mouse Ensembl gene ID. ClinVar ID and COSMIC ID carry no species selector: both are human variant databases, and both resolve against the human reference.Homology-Directed Repair (HDR) Donor TemplatesOn this workflow the donor columns belong to finder mode, which is where guides are enumerated.There is no HDR mode switch: the input format decides. When the input format specifies an edit — an unedited/edited sequence pair, a ClinVar ID or a COSMIC ID — every candidate guide is also reported with what it needs for HDR. The other six formats identify the target without specifying an edit, so no donor is designed from them and the donor columns are absent rather than blank.Cas12a does not cut bluntly. It leaves a staggered break distal to the PAM, so both boundaries are reported — Cut site (1-based in the target sequence) and Cut site (opposite strand) — and a donor's arms are measured from the middle of the wound rather than from a single position.Each candidate carries these columns, in the results table and in the downloadable files alike:

Cut-to-edit distance (nt) — how far the break sits from the base being changed. Candidates are ranked by it (HDR donor rank in the target), because repair efficiency falls away with distance; candidates at the same distance keep the order the model listed them in.

Donor (sense) / Donor (antisense) — the single-stranded donor carrying the edit, in the orientation(s) you asked for, ready to order.

Donor length (nt), Left arm (nt), Right arm (nt), Requested arms (nt) — arms are measured outward from the union of the cut site and the edit, not from either alone, so that a guide cutting some distance away still spans both. The realised arms therefore differ from the requested pair whenever the cut sits outside the edit, and both are reported.

Indel length not a multiple of 3 — a length test on the edit itself. It is deliberately not called frame-disrupting: the donor is built from a sequence window, so whether the position is coding at all is not knowable here.

Blocking mutation — off by default. When enabled, a change that stops the nuclease re-cutting the repaired allele.

Donor note — what shaped this donor: the staggered break the arms are measured across, why a blocking mutation was or was not made, and by how much the arms were extended outward past the requested lengths so that the donor spans the cut as well as the edit. Where a row carries no donor, the note gives the reason in its place — arms that do not fit inside the supplied window, naming the extra sequence or the shorter arm that would make them fit, or a window that could not be tied to exactly one input record.

What you can set: homology arm lengths (100/100 nt by default, 50/50, or a custom pair between 25 and 1000 nt), the donor strand, whether to add a blocking mutation, and the donor template type — a chemically synthesized single-stranded oligodeoxynucleotide (ssODN) by default, with the donor note saying when a template outgrows standard oligo synthesis and should be ordered as a long single-stranded DNA (ssDNA) fragment instead; long ssDNA for those longer templates; or a double-stranded DNA (dsDNA) duplex, which arrives as one Duplex donor (dsDNA) column instead of the strand pair. The guide sequence is deliberately not yours to specify — the guides are this model's own ranked candidates, and a donor is built for each of them.Guide Cloning OligosCloning Vector decides whether the results carry an ordering-ready oligo pair. Two Cas12a vectors are offered — Cas12a / pY108 or pU6-crRNA (BsmBI), the default, and Cas12a / pRDA_052 Broad GPP (BsmBI) — and every candidate then gains Oligo (forward) and Oligo (reverse), in the results table and in the downloadable files alike: the two strands that anneal into the digested vector, each presenting the 5' overhang that vector needs. A Cas12a crRNA takes no initiating G, so the protospacer is carried as it stands, and it is the full 23 nt protospacer the model scores rather than any shorter view of it.Setting the vector to None (No Oligo Output) leaves both columns out rather than blank. The four overhang fields — Forward 5′, Reverse 5′, Forward 3′ and Reverse 3′ — override what the vector supplies; left blank, each takes the vector's own. A guide carrying anything outside A, C, G and T gives an empty pair, because there is nothing orderable to write.Off-Target ScoreThe Off-target score (higher = less specific) column is a mismatch-tolerance score, not a count of sites in a genome. It asks a narrower question than its name suggests: for this guide, how much activity would survive a single mismatch, averaged over every single-mismatch variant of the protospacer. A guide that tolerates mismatches well is a guide that is more likely to cut somewhere it was not aimed at, which is why a higher number is the worse one.It is computed from the Cas12a substitution table (DeWeirdt et al. 2021, scored by the scheme of Doench, Fusi et al. 2016), so it needs no genome and runs in the time the on-target prediction already takes. It cannot tell you where an off-target is, and it does not know whether any of those variants occurs in your genome at all.For that, set Genome-wide off-target search to Human or Mouse. That search adds one Genome-wide specificity (higher = more specific) column, which runs the other way round: there, higher is better. Every site it finds is condensed into that single number; the individual loci are not listed, so no chromosome, coordinate or strand for them reaches the table. The two are different measurements and both may appear. The sites that search finds are scored on the same table, so the two columns differ only in the question they answer -- one guide against every single-mismatch variant of itself, one guide against the sites a genome really holds -- and never in how a mismatch is weighed.ResultsClick any row in the output table to update the detail panel. Result_Summary

PAM regions are highlighted in red.

Score distribution and top-score summaries are provided.

A Download Results button is available at the bottom of the page.

Sample OutputsThe following example interactive outputs are available for reference:Single guideBatch scoringGuide finder, FASTAGuide finder, gene symbolGuide finder, ClinVar variantData and LicensingCOSMIC. COSMIC is a product of Genome Research Limited (Wellcome Sanger Institute). It is free for academic and non-commercial use; commercial use requires a COSMIC licence. Lookups here resolve a single user-typed identifier — no COSMIC data is listed, browsable or served in bulk.ContactIf you have any questions, bug reports, or suggestions, please do not hesitate to contact us through the Support page.

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