Evaluation_of_whole_genome_amplification_and_bioinformatic_methods_for_the_characterization_of_Leishmania_genomes_at_a_single_cell_level-sc-3568
Evaluation_of_whole_genome_amplification_and_bioinformatic_methods_for_the_characterization_of_Leishmania_genomes_at_a_single_cell_level
Here, we report a pilot study paving the way for further single cell genomics studies in Leishmania. First, the performances of two commercially available kits for Whole Genome Amplification (WGA), PicoPlex and RepliG was compared on small amounts of Leishmania donovani DNA, testing their ability to preserve specific genetic variations, including aneuploidy levels and SNPs. We show here that the choice of WGA method should be determined by the planned downstream genetic analysis, Picoplex and RepliG performing better for aneuploidy and SNP calling, respectively. This comparison allowed us to evaluate and optimize corresponding bio-informatic methods. As PicoPlex was shown to be the preferred method for studying single cell aneuploidy, this method was applied in a second step, on single cells of L. braziliensis, which were sorted by fluorescence activated cell sorting (FACS). Even sequencing depth was achieved in 28 single cells, allowing accurate somy estimation. A dominant karyotype with three aneuploid chromosomes was observed in 25 cells, while two different minor karyotypes were observed in the other cells. Our method thus allowed the detection of aneuploidy mosaicism, and provides a solid basis which can be further refined to concur with higher-throughput single cell genomic methods. This data has been described in the following article doi: https://doi.org/10.1101/2020.02.20.957621  and its further analysis can be freely submitted for publication. For information on the proper use of data shared by the Wellcome Trust Sanger Institute (including information on acknowledgement), please see http://www.sanger.ac.uk/datasharing/
Illumina sequencing and enrichment of the whole L.donovani genome from pooled and single cells.