---
title: Interpreting CRISPR outputs
description: >-
  Guide RNA tables, on-target scores, off-target site reports, and CRISPR screen
  gene rankings.
tags:
  - crispr
  - outputs
---

These tools return ranked guide RNA tables with on-target scores, off-target site reports with mismatch counts, and screen gene rankings — letting you pick a high-efficiency, low-risk guide for the bench or shortlist screen hits for follow-up.

## Interpreting CRISPR outputs

## Guide RNA Design output

| Field | Type | Description |
|-------|------|-------------|
| `gene` | string | Target gene symbol |
| `organism` | string | Target organism |
| `pam` | string | PAM sequence used |
| `total_sites` | number | Total PAM sites found in the target region |
| `returned_count` | number | Number of guides returned (after filtering) |
| `guides` | array | Ranked guide RNA candidates |

### Guide fields

| Field | Type | Description |
|-------|------|-------------|
| `rank` | number | Rank by on-target score (1 = best) |
| `sequence` | string | 20-nt guide RNA sequence (DNA) |
| `pam_sequence` | string | PAM sequence at the target site |
| `strand` | string | Target strand (`+` or `-`) |
| `chromosome` | string | Chromosome name |
| `position` | number | Genomic start position |
| `exon` | string | Exon name if in a coding exon (e.g., `exon3`) |
| `gc_content` | number | GC percentage of the guide (0-100) |
| `on_target_score` | number | Predicted cutting efficiency (0-1) |

### On-target score interpretation

| Score Range | Interpretation |
|-------------|----------------|
| 0.7 - 1.0 | High predicted activity; recommended |
| 0.5 - 0.7 | Moderate activity; acceptable for most experiments |
| 0.3 - 0.5 | Low activity; may require higher MOI or alternative delivery |
| < 0.3 | Poor predicted activity; not recommended |

### GC content guidelines

| GC% | Interpretation |
|-----|----------------|
| 40 - 60% | Optimal range for most applications |
| 30 - 40% or 60 - 70% | Acceptable; may have slightly reduced activity |
| < 30% or > 70% | Suboptimal; higher risk of secondary structure or stability issues |

## Off-Target Analysis output

| Field | Type | Description |
|-------|------|-------------|
| `guide` | string | Query guide sequence |
| `genome` | string | Reference genome used |
| `pam` | string | PAM filter applied |
| `total_offtargets` | number | Total off-target sites found |
| `offtargets_by_mismatch` | object | Count of sites per mismatch level |
| `sites` | array | Off-target site details |

### Off-target site fields

| Field | Type | Description |
|-------|------|-------------|
| `sequence` | string | Off-target genomic sequence |
| `mismatches` | number | Number of mismatches relative to the guide |
| `mismatch_positions` | number[] | Positions where mismatches occur (1-indexed from PAM-proximal end) |
| `chromosome` | string | Chromosome |
| `position` | number | Genomic position |
| `strand` | string | Strand (`+` or `-`) |
| `gene` | string | Nearest gene symbol (or `--` if intergenic) |
| `in_cds` | boolean | Whether the site falls within a coding sequence |
| `in_exon` | boolean | Whether the site falls within any exon |

### Off-target risk assessment

| Mismatch Count | 0 | 1 | 2 | 3 |
|----------------|---|---|---|---|
| Expected | 1 (on-target) | 0-2 | 0-20 | 0-200 |
| Concern level | N/A | High | Moderate | Low |

A guide is considered low-risk when:
- Zero 1-mismatch off-target sites outside the intended target
- Fewer than 5 sites at 2 mismatches in coding regions
- No off-target sites in known essential genes

## Screen Analysis output

| Field | Type | Description |
|-------|------|-------------|
| `analysis_type` | string | `"mageck"` |
| `direction` | string | Analysis direction (`negative`, `positive`, or `both`) |
| `total_genes` | number | Total genes analyzed |
| `significant_genes` | number | Genes passing FDR threshold |
| `fdr_threshold` | number | FDR cutoff used |
| `gene_rankings` | array | Gene-level results sorted by significance |

### Gene ranking fields

| Field | Type | Description |
|-------|------|-------------|
| `rank` | number | Gene rank (1 = most significant) |
| `gene` | string | Gene symbol |
| `num_guides` | number | Number of guides targeting this gene |
| `guides_significant` | number | Guides showing consistent effect |
| `rra_score` | number | Robust Rank Aggregation score (lower = more significant) |
| `p_value` | number | Gene-level p-value |
| `fdr` | number | Benjamini-Hochberg adjusted p-value |
| `lfc` | number | Median log2 fold change across guides |
| `significant` | boolean | Whether the gene passes FDR threshold |

### RRA score interpretation

| RRA Score | Interpretation |
|-----------|----------------|
| < 1e-5 | Very strong hit; high confidence |
| 1e-5 to 1e-3 | Strong hit; likely true positive |
| 1e-3 to 0.01 | Moderate hit; requires validation |
| > 0.01 | Weak or non-significant |

### Log2 fold change interpretation

| Direction | LFC < 0 | LFC > 0 |
|-----------|---------|---------|
| Negative selection | Guide depletion (essential gene) | Guide enrichment (not depleted) |
| Positive selection | Guide depletion (not enriched) | Guide enrichment (resistance gene) |

## Read next