CFD updates

Cargo.toml

@@ -13,3 +13,4 @@ lib_wfa2 = { git = "https://github.com/AndreaGuarracino/lib_wfa2", rev = "c608c4
 rand = { version = "0.9.0", features = ["small_rng"] }
 rayon = "1.10.0"
 flate2 = "1.1.0"
+lazy_static = "1.4"

README.md

@@ -13,7 +13,7 @@ CRISPRapido is a reference-free tool for comprehensive detection of CRISPR off-t
 - Automatic reverse complement scanning
 - PAF-format output compatible with downstream analysis tools
 - Multi-threaded processing for improved performance
-
+- CFD (Cutting Frequency Determination) scoring for off-targets
 ## Installation
 
 You need to build `WFA2-lib` first, which is a submodule of this repository. To do so, run:
@@ -50,14 +50,14 @@ env -i bash -c 'WFA2LIB_PATH="./WFA2-lib" PATH=/usr/local/bin:/usr/bin:/bin ~/.c
 ## Usage
 
 ```bash
-crisprapido -r <reference.fa> -g <guide_sequence> [OPTIONS]
+crisprapido -r <reference.fa> -g <guide_sequence> -p <pam_sequence> [OPTIONS]
 ```
 
 ### Required Arguments
 
 - `-r, --reference <FILE>`: Input reference FASTA file (supports .fa and .fa.gz)
 - `-g, --guide <SEQUENCE>`: Guide RNA sequence (without PAM)
-
+- `-p, --pam <SEQUENCE>` : PAM sequence for CFD
 ### Optional Arguments
 
 - `-m, --max-mismatches <NUM>`: Maximum number of mismatches allowed (default: 4)
@@ -95,11 +95,26 @@ Additionally, CRISPRapido includes these custom tags:
 | `ng:i` | Number of gaps (indels) |
 | `bs:i` | Biggest gap size in bases |
 | `cg:Z` | CIGAR string representing alignment details |
+| `cf:f` | CFD score
+
+
+### CFD Score
 
+The Cutting Frequency Determination (CFD) score estimates the likelihood of a guide RNA cutting at an off-target site.
+The score ranges from 0.0 to 1.0,  taking into account:
+
+- Position-specific mismatch penalties
+- PAM sequence efficiency
+- Bulge and gap effects
+
+This implementation requires two data files:
+
+- `mismatch_scores.txt` : Position-specific mismatch penalties
+- `pam_scores.txt` : Efficiency scores for different PAM sequences
 ### Example Output
 
 ```
-Guide   20      0       20      +       chr1    248956422       10050   10070   19      21      255     as:i:6  nm:i:1  ng:i:0  bs:i:0  cg:Z:19=1X
+Guide   20      0       20      +       chr1    248956422       10050   10070   19      21      255     as:i:6  nm:i:1  ng:i:0  bs:i:0  cg:Z:19=1X  cf:f:0.0549
 ```
 
 This indicates:
@@ -117,7 +132,7 @@ For more details on the PAF format, see the [official specification](https://git
 ## Example
 
 ```bash
-crisprapido -r genome.fa -g ATCGATCGATCG -m 3 -b 1 -z 2
+crisprapido -r genome.fa -g ATCGATCGATCG -p GG -m 3 -b 1 -z 2
 ```
 
 ## Testing
@@ -144,3 +159,4 @@ See LICENSE file
 ## Citation
 
 Stay tuned!
+