fools

FISH tools (fools) to reproducibly generate FISH probes with PaintShop and oligominer

Installation

pip install git+https://github.com/adrtsc/fools.git

Specification of metadata file for PaintShop probes

The metadata file contains all the information about the probe color, primer pair to use and the oligo pool the probes are associated with. It has to be submitted as .csv file with at least the following columns:

gene_name	color	primer_pair	pool_name
HSP60	red	1	HSP60-TP53
TP53	far-red	2	HSP60-TP53

• The "gene_name" column contains the gene symbol of the target gene.
• The "color" column can have the values green, red and far-red. It defines which barcode will be appended to the probe and therefore which color the probe will be in the end.
• The "primer_pair" can have the values 1 or 2. The column defines which primer sequence will be appended. The primer sequences are used for the clean-up protocol of the probes.
• The "pool_name" column defines the name of the pool associated to the probes of that gene in the IDT order form that is generated by the scripts.

Specification of metadata file for oligominer probes

We use oligominer mainly to generate intronic FISH probes. Therefore the metadata file needs to include all columns as above, but also chromosome coordinates of interest:

gene_name	color	primer_pair	pool_name	start	end
HSP60	red	1	HSP60-TP53	11020230	11023230
TP53	far-red	2	HSP60-TP53	919239	921239

• The "start" column defines the start coordinate of the region of interest
• The "end" column defines the end coordrinate of the region of interest

The chromosome number and the coding strand of the gene do not have to be included in the metadata file, they will be inferred from the annotation file.

Generating a multi-input fasta file for oligominer:

generate_fasta --genome_path /path/to/genome.fasta --annotation_path /path/to/annotations.gtf --metadata_path /path/to/metadata.csv

Generating an IDT order file from paintshop output:

generate_probes --probe_path /path/to/paintshop/result.txt --metadata_path /path/to/metadata.csv

Generating an IDT order file from oligominer output:

generate_probes --probe_path /path/to/oligominer/result.csv --metadata_path /path/to/metadata.csv --probe_design oligominer