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PRBB-CRG Sessions Jan-Philipp Junker

PRBB-CRG Sessions Jan-Philipp Junker

13/09/2019
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PRBB-CRG Sessions Jan-Philipp Junker

MARIE CURIE

13/09/201912:00MARIE CURIEPRBB-CRG SessionsJan-Philipp JunkerMax-Delbrück Center for Molecular Medicine (MDC), Berlin, DE"Lineage tracing on the organism-wide level"Host: Arnau Sebé Pedrós (CRG)Abstract:A key goal of developmental biology is to understand how a single cell transforms into a full-grown organism consisting of many cells. Although impressive progress has been made in lineage tracing using imaging approaches, analysis of vertebrate lineage trees has mostly been limited to relatively small subsets of cells. Here we present LINNAEUS, a strategy for massively parallel lineage tracing on the single cell level. We exploit the fact that Cas9-induced generation of double-strand breaks leads to formation of short insertions or deletions (“genetic scars”) that are variable in their length and position. We demonstrate that these genetic scars have very high complexity, making them ideal cellular barcodes for lineage analysis. Reading out scar sequences in expressed transgenes by single-cell RNA-seq enables us to simultaneously measure lineage relationships and transcriptome profiles in thousands of single cells. We introduce novel computational methods that address the major challenges of CRISPR/Cas9-based lineage tracing, such as preferential creation of specific scar sequences and stochastic dropout events. We use LINNAEUS to reconstruct developmental lineage trees in zebrafish at larval stages as well as in dissociated organs of adult fish, and we demonstrate the potential of our approach for studying organ regeneration upon injury. LINNAEUS allows scaling up lineage analysis to the level of the whole organism, and it will enable systematic studies of lineage variability and robustness of lineage trees to perturbation.