evvie_crispr_scripts

guide_assignment_9.26.23_w_tar_and_reformat_perturb.sh is a bash script designed to take the output of a 10x cell ranger analysis of a CRISPRi dataset, compute guide assignmnet, produce guide assignment QCs, and create output files for downstream processes.

The script can be run on a slurm-based system as follows:

sbatch guide_assignment_9.26.23_w_tar_and_reformat_perturb.sh SAMPLE_NAME PATH_to_10x_DIR_ENDING_IN/ MIN_GUIDE_UMI_THRESHOLD Y_N_USE_10x_GUIDE_CALLS Y_N_Testing

For example:

sbatch guide_assignment_9.26.23_w_tar_and_reformat_perturb.sh 230327_Encode_Tap_MOI5_sample7_novaseq_and_qc_seq /oak/stanford/groups/engreitz/Users/ejagoda/230327_Encode_K562_Tap_seq_full_seq/230327_Encode_Tap_MOI5_sample7_novaseq_and_qc_seq/ 2 yes no

**The script will do as follows:*,

1. unzip the 10x cell ranger outputs crispr_analysis.tar.gz and filtered_feature_bc_matrix.tar.gz
2. call the script all_upstream_of_mast_read_edits9.22.23_double_check.R which computes guide assignments and creates the output files described below. If the Y_N_USE_10x_GUIDE_CALLS flag is set to "no", guide assignmnets will be made uniformly based on the minimum umi trehshold which can be adjusted in the MIN_GUIDE_UMI_THRESHOLD theshold flag (default = 2). If the Y_N_USE_10x_GUIDE_CALLS flag is set to "yes", guide asssignments will be primarly based on the 10x calls, unless the guide threshold is less than the minimum threshold in which case the minimum threshold will be used. To strictly follow the 10x guide calls, set the guide umi threshold to 1.
3. modify the header of the "_perturb_status.txt" to make it useable for the MAST snakemake

output files from step 2

1. _protospacer_calls_per_cell.csv - final guide calls per cell formatted in the cellranger format
2. _guides_per_cell.txt - CBC x #guides assigned
3. _per_guide_summary_table.txt - guide - number cells assigned - umi threshold used
4. _perturb_status.txt - binary matrix guide assignments in the form of CBC x guide
5. _Cells_per_guide.png - histogram of the number of cells assigned to each guide
6. _Guides_per_cell.png - histogram of the number of guides assigned to each cell
7. _umi_threshold_hist_log10.png - hisogram of number of minimum guide umis needed to assign each guide to a cell
8. _assinged_vs_not_assinged_umis_cdf.png - per cell CDF plot of ratio of guide umis per cell that were attributed to an assigned guide vs a non-assigned guide (ambient rna)
9. _assinged_vs_not_assinged_umis_boxplot.png - boxplot of ratio of guide umis per cell that were attributed to an assigned guide vs a non-assigned guide (ambient rna)