TIDE (Tracking of Indels by DEcomposition) quantifies the outcome of a CRISPR/Cas9, TALEN or ZFN experiment from a single Sanger sequencing reaction. The app compares the chromatogram of an edited cell population with a control trace and decomposes the mixed signal downstream of the cut site into the individual insertions and deletions that produced it.
The result is an overall editing efficiency percentage, the frequency of every indel in the spectrum, and an R² value describing how well the decomposition explains the observed trace. No next-generation sequencing, library preparation or bioinformatics support is required.
A CRISPR edit produces a heterogeneous population of cells, each carrying a different indel. When this population is sequenced by Sanger capillary electrophoresis, the resulting chromatogram is a superposition of every trace weighted by its frequency. TIDE reconstructs those frequencies by non-linear least-squares decomposition, turning a single mixed trace into a quantitative indel spectrum.
The algorithm first aligns the control and edited traces, locates the expected cut site from the 20-nucleotide guide RNA sequence, and generates a library of theoretical chromatograms for every insertion and deletion within the selected size range. It then fits the observed edited trace as a linear combination of these theoretical traces, constrained so that the estimated frequencies sum to one and remain non-negative.
The result is a precise editing efficiency percentage, the relative frequency of each indel size, and an R² value that reports how much of the post-cut variance the model explains. Because the method uses only two Sanger reactions per sample, it offers a fast, low-cost alternative to amplicon deep sequencing for routine indel quantification and CRISPR screening.
Step 1
Provide the .ab1 or .scf file of a non-edited sample amplified with the same primer pair. This trace defines the unedited reference sequence.
Step 2
Provide the Sanger trace of the PCR amplicon from your edited population. Both traces should cover at least 100 bases before and several hundred bases after the expected break site.
Step 3
Paste the 20-nucleotide protospacer without the PAM. TIDE uses it to locate the expected cut site and to set the decomposition window.
Step 4
The default window covers the region after the cut site with reliable sequence quality. The indel size range determines how large the detected insertions and deletions may be.
Step 5
Within seconds you receive the total editing efficiency, a bar chart of the indel spectrum with p-values per indel size, the aligned sequences and quality control statistics you can export for your records.
Frequencies for every insertion and deletion size within the selected range, each with a significance value against the control trace.
Two capillary sequencing reactions per sample are sufficient. Most providers deliver results within one working day.
R² of the decomposition, aberrant sequence signal before the cut site and trace quality warnings help you judge whether a result is trustworthy.
The algorithm is described in Brinkman et al., Nucleic Acids Research 2014, and has been cited in thousands of gene editing studies.
TIDE requires two Sanger sequencing trace files (.ab1 or .scf), one from a control sample and one from the edited population, together with the 20-nucleotide guide RNA sequence.
Published comparisons show that TIDE editing efficiencies correlate closely with amplicon deep sequencing for indels within the selected size range, at a fraction of the cost and turnaround time.
TIDE reliably decomposes insertions and deletions up to approximately 50 nucleotides. Larger rearrangements are not resolved and appear as unexplained signal, visible in the R² value.
For homology-directed repair and other template-based edits, the related TIDER method extends the decomposition with a reference template trace.
The research version of the app is free for non-profit research use. A validated, audit-ready TIDE GxP tier is available for regulated environments.