Published On: Thu, Aug 9th, 2018

Genetic barcodes can record every cell’s history in real time

The researchers implemented the hgRNA-Cas9 system in mice by creating a “founder mouse” that had 60 different hgRNA sequences scattered throughout its genome. They then crossed the founder mouse with mice that expressed the Cas9 protein, producing zygotes whose hgRNA sequences started being cut and mutated shortly after fertilization.

“Starting with the zygote and continuing through all of its progeny, every time a cell divides there’s a chance that its daughter cells’ hgRNAs will mutate,” explained first author Reza Kalhor, a postdoctoral research fellow at the Wyss Institute and HMS. “In each generation, all the cells acquire their own unique mutations in addition to the ones they inherit from their mother cell, so we can trace how closely related different cells are by comparing which mutations they have.”

Each hgRNA can produce hundreds of mutant alleles; collectively, they can generate a unique barcode that contains the full developmental lineage of each of the approximately 10 billion cells in an adult mouse.

The ability to continuously record cells’ development also allowed the researchers to resolve a longstanding question regarding the embryonic brain: Does it distinguish its front from its back end first, or its left from its right side? By comparing the hgRNA mutation barcodes present in cells taken from different parts of two mice’s brains, they found that neurons from the left side of each brain region were more closely related to neurons from the right side of the same region than to neurons from the left side of neighboring regions. This result suggested that front-back brain patterning emerges before left-right patterning in central nervous system development.

“This method allows us to take the final developmental stage of a model organism and from there reconstruct a full lineage tree all the way back to its single-cell stage. It’s an ambitious goal that will certainly take many labs several years to realize, but this paper represents an important step in getting there,” said Church.

The researchers are now focusing on improving their readout techniques so that they can analyze the barcodes of individual cells and reconstruct the lineage tree that has been recorded.

“Being able to record cells continuously over time is a huge milestone in developmental biology that promises to exponentially increase our understanding of the process by which a single cell grows to form to an adult animal and, if applied to disease models, it could provide entirely new insights into how diseases, such as cancer, emerge,” said Donald Ingber, director of the Wyss Institute. Ingber is also the Judah Folkman Professor of Vascular Biology at HMS and the vascular biology program at Boston Children’s Hospital, and professor of bioengineering at Harvard’s John A. Paulson School of Engineering and Applied Sciences.

Additional authors of the paper include Kian Kalhor from Sharif University of Technology in Tehran, Iran; Leo Mejia from HMS; Kathleen Leeper and Amanda Graveline from the Wyss Institute; and Prashant Mali, associate professor at the University of California, San Diego.

This research was supported by the National Institutes of Health and the Intelligence Advanced Research Projects Activity.

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Science & Technology – Harvard Gazette

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Genetic barcodes can record every cell’s history in real time