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Functional Screening in L. mexicana Understanding the genetics of drug response in the protozoan Leishmania is critical for treatment strategies but is hindered by the parasite’s lack of RNAi and non-homologous end-joining. In Arias et al., BioRxiv 2025, we addressed this using CRISPR/Cas9 cytosine base editing for genome-wide loss-of-function screening in L. mexicana. The resulting datasets, accessible here at LeishBASEeditDB.net, revealed numerous novel resistance and sensitivity biomarkers across five compounds: SbIII, miltefosine, amphotericin B, pentamidine, and the experimental arylmethylaminosteroid 1c. A library consisting of 38,422 protein-targeting CBE sgRNA expression cassettes was generated, with 1 to 6 sgRNAs per gene (~5.2 on average). These sgRNAs were designed to introduce STOP codons within the first 50% of coding sequences in 94% of all protein-coding ORFs. Additionally, 1,000 non-targeting control sgRNAs were included. The entire library was integrated into the 18S rRNA SSU locus via Cas12a-mediated recombination, at a rate of 500 parasites per sgRNA (following the method described in Engstler and Beneke, eLife 2023 and Herrmann May et al., eLife 2025). Expressing one sgRNA per cell, this enabled the functional assessment of 7,378 unique and 340 multi-copy or isoform genes in L. mexicana. This website provides access to the resulting data generated using a MAGeCK analysis pipeline. The displayed data includes the average fold change (FC) of all sgRNAs per gene across three independent replicates. A p-value is calculated based on the performance of all sgRNAs per gene, and the average across three independent replicates is shown. The data for each individual screen can be accessed on separate pages and genes can be selected using the search form on the left. Please click the icons below to navigate between pages. You can download a '.csv' file with the complete data (individual sgRNAs or sgRNA-sets) for each screen here: |
The sgRNA viewer and triplicate deviation are enabled for <300 Gene IDs. For <10 Gene IDs, sgRNAs and genes are color-coded. |
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Pentamidine CRISPR Screening Data The sgRNA library was transfected into L. mexicana promastigotes and cultured for 144 hours before treatment with Pentamidine. Libraries were treated for 48 hours, washed, and surviving cells were allowed to recover over 72 hours. DNA from the resulting population was isolated at this time point (recovery 1) and after an additional 48 hours of sub-culture (recovery 2). In parallel, a non-treated library was cultured, and DNA was isolated. Both recoveries and controls were subjected to Illumina sequencing to determine sgRNA abundances. The data compares sgRNA abundance in recovery 1 and recovery 2 with their respective control samples. Click on data points to get to TritrypDB gene page! For "Gene Charts," the median p-value and median fold change for all sgRNAs per gene is displayed. Since there are on average five sgRNAs per gene, non-targets are randomly grouped into sets of five. For "sgRNA Charts," each sgRNA or non-target control is shown individually. |
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Gene - 264h vs PTM Recovery 1 |
Gene - 312h vs PTM Recovery 2 |
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Select other CRISPR screening data by clicking the icons below! |
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Website designed and maintained by Tom Beneke (tobeneke@gmail.com or tom.beneke@uni-wuerzburg.de). Last updated on 17th June 2025.