A novel study from researchers at Karolinska Institutet shows that by using so called random sequence labels, working as barcodes for tracing genetically altered cells, during CRISPR/Cas9 gene knock-outs it is possible to generate more comprehensive data from a smaller number of cells. The innovation has led to the SciLifeLab High Throughput Genome Engineering facility implementing barcodes in all future screening projects.
CRISPR/Cas9 gene knock-out uses guide-RNAs to target the nuclease Cas9 to defined regions in the genome of cells. Cas9 then cuts the genomic DNA precisely at the position specified by the guide-RNA, and erroneous repair leads to gene disruption. In CRISPR/Cas9 genetic screens, a whole library of guide sequences is integrated into the genome of target cells such that each cell expresses a single guide RNA against one specific gene. Cells with characteristics of interest are selected from this mutant cell population and analyzed. To obtain meaningful results, such screens typically need to be done with several hundred cells per guide sequence.
The Scilifelab High Throughput Genome Engineering (HTGE) facility, in collaboration with Jussi Taipale’s research team, have now barcoded guide-sequences with so called random sequence labels and demonstrated that the approach adds value to CRISPR/Cas9 screens.
“The innovation allows tracking of single cell lineages throughout a screen, which creates a much richer dataset from a smaller number of cells.”, says Bernhard Schmierer, first author of the paper and Facility Director of the HTGE facility. ” The novel approach substantially improves accuracy and reliability of CRISPR/Cas9 screens, while incurring little extra cost. Our facility will use barcoded guide libraries in all future screening projects.”
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