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<jats:title>Abstract</jats:title><jats:p>The number of fully sequenced genomes increases steadily but the function of many genes remains unstudied. To accelerate dissection of gene function in <jats:italic>Leishmania</jats:italic> spp. and other kinetoplastids we developed previously a streamlined pipeline for CRISPR-Cas9 gene editing, which we termed LeishGEdit [1]. To facilitate high-throughput mutant screens we have adapted this pipeline by barcoding mutants with unique 17-nucleotide barcodes, allowing loss-of-function screens in mixed populations [2]. Here we present primer design and analysis tools that facilitate these bar-seq strategies. We have developed a standalone easy-to-use pipeline to design CRISPR primers suitable for the LeishGEdit toolbox for any given genome and have generated a list of 14,995 barcodes. Barcodes and oligos are now accessible through our website <jats:ext-link xmlns:xlink="http://www.w3.org/1999/xlink" ext-link-type="uri" xlink:href="http://www.leishgedit.net">www.leishgedit.net</jats:ext-link> allowing to pursue bar-seq experiments in all currently available TriTrypDB genomes (release 41). This will streamline CRISPR bar-seq assays in kinetoplastids, enabling pooled mutant screens across the community.</jats:p><jats:sec><jats:title>Highlights</jats:title><jats:list list-type="bullet"><jats:list-item><jats:p>Developing tools for pooled bar-seq mutant screens across the kinetoplastid community</jats:p></jats:list-item><jats:list-item><jats:p>Development of a standalone script to design primers suitable for the LeishGEdit toolbox</jats:p></jats:list-item><jats:list-item><jats:p>Generation of 14,995 barcodes that can be used for bar-seq strategies in kinetoplastids</jats:p></jats:list-item><jats:list-item><jats:p>Bar-seq primers for all TriTrypDB genomes (release 41) can be obtained from <jats:ext-link xmlns:xlink="http://www.w3.org/1999/xlink" ext-link-type="uri" xlink:href="http://www.leishgedit.net">www.leishgedit.net</jats:ext-link></jats:p></jats:list-item></jats:list></jats:sec>

Original publication

DOI

10.1101/2020.03.19.998856

Type

Journal article

Publisher

Cold Spring Harbor Laboratory

Publication Date

20/03/2020