High-throughput dense reconstruction of cell lineages

Abstract

The first meeting exclusively dedicated to the ‘High-throughput dense reconstruction of cell lineages' took place at Janelia Research Campus (Howard Hughes Medical Institute) from 14 to 18 April 2019. Organized by Tzumin Lee, Connie Cepko, Jorge Garcia-Marques and Isabel Espinosa-Medina, this meeting echoed the recent eruption of new tools that allow the reconstruction of lineages based on the phylogenetic analysis of DNA mutations induced during development. Combined with single-cell RNA sequencing, these tools promise to solve the lineage of complex model organisms at single-cell resolution. Here, we compile the conference consensus on the technological and computational challenges emerging from the use of the new strategies, as well as potential solutions.

Imaging cell lineage with a synthetic digital recording system

“…Concatenating multiple trits in a compact array and integrating it at a safe harbor locus ( 35 – 37 ) increases the amount of memory while facilitating germline transmissibility ( 38 ). Edit strategies that rely on double-stranded breaks and endogenous DNA repair machinery are prone to information loss in arrays through inter-unit deletions ( 27 , 39 ). By contrast, integrases allow recombinational isolation between distinct trits in the same array.…”

Dense reconstruction of elongated cell lineages: overcoming suboptimum lineage encoding and sparse cell sampling

“…However inter-target deletions can occur if Cas9 targets are on the same chromosome. It has been shown that frequent, large, inter-target deletions diminish the ability to accurately perform phylogenetic analyses [30,18]. Inter-target deletions would be particularly problematic if the Cas9 targets are placed in an array, as coding units or previous edits can easily be lost [10].…”

“…The CRISPR-based method creates evolving genetic barcodes via a cumulative process of CRISPR editing. Cas9, in combination with a gRNA, modifies specific predefined sequences in the cell (for more details see [18,19,20]). This editing process (encoding) produces independent edits in different cells.…”

Single-cell lineage tracing approaches in hematology research: technical considerations

“…The labeling of cells in the native setting poses many challenges, across multiple disciplines and tissues [52].…”