Zebrafish, a small, fast-growing creature that shares many of the same genes as humans, can study a wide range of questions, from how organisms develop to how the nervous system drives behavior. It is useful for many biologists who find it very suitable. A new technology, now called MIC-Drop, developed by health scientists at the University of Utah makes fish even more powerful for large-scale genetic research.
MIC-Drop, led by chemical biologist Dr. Randall Peterson (University of Utah, Dean of the University of Pharmaceutical Sciences), allows researchers to efficiently introduce a CRISPR gene editing system into zebrafish to function hundreds of genes. Can be evaluated quickly. In one experiment. This advance is the first time that screening using robust Nobel Prize-winning CRISPR technology has become possible in any animal model. Already, Peterson’s team has used MIC-Drop to identify some genes that are essential for the healthy development and function of the heart.Their methods and discoveries were reported in the journal on August 19, 2021. Chemistry..
The CRISPR system is a programmable method for modifying DNA. To use it, researchers introduce a DNA-cleaving enzyme (usually an enzyme called Cas9) into cells and attach an RNA guide to the enzyme to indicate where it is cut. This can be the first step in altering the sequence of a gene, or simply blocking the gene.
This method has made genetic editing of zebrafish and other experimental organisms faster, cheaper and more accurate. However, according to Peterson, it was difficult to scale up to study more than a few genes at a time.To inactivate a single gene in Zebrafish embryo, Researchers prepare a guide RNA that targets the gene, mix it with the Cas9 enzyme, load the solution into a needle, and inject a carefully calibrated amount of solution into the embryo. If you want to inactivate another gene in another embryo, you need to load a new Cas9 / guide RNA solution into a new needle. “This process has always focused on one gene or one modification at a time,” says Peterson. “That is, if you want to execute 100 genes, it’s 100 times more work.”
MIC-Drop, an abbreviation for Multiplexed Intermixed CRISPR Droplets, solves this problem by packaging the components of the CRISPR system into fine oil-covered droplets that can be mixed without confusion. I will solve it. To set up screening for many genes using MIC-Drop, researchers begin by creating a library of guide RNAs. Each guide RNA is packaged in its own droplet with Cas9 enzyme. To track the target gene, every drop also contains a DNA barcode that identifies its contents.
The team fine-tuned the chemistry of the droplets to keep them stable and discrete. Therefore, droplets designed to target different genes can be mixed and loaded onto the same needle. Under the microscope, the MIC-Drop user injects a single drop into a zebrafish embryo, then moves to the next embryo and injects the next drop.This process can be repeated hundreds of times and delivers a single packet of CRISPR components to each embryo, so the system in all embryos Single gene.. Second, it’s up to the researcher to monitor the animals for potential effects.
Previously, setting up a CRISPR screen for hundreds of genes in zebrafish took days and required hundreds of needles, and developed MIC-Drop’s packaging technology and barcodes. And optimized postdoctoral fellow Dr. Saba Parvez said. system. “Now we can streamline the process to a single user and run it in a span of hours,” he says.
To demonstrate the potential of MIC-Drop, Parvez et al., H. Joseph Yost, Ph.D., Calum MacRae, MD, Ph.D., colleagues at the U of U Health at Harvard Medical School. , And Jing-Ruey Joanna Yeh, Ph. .D. , Massachusetts General Hospital, tested 188 different zebrafish genes for their potential role in heart development. After creating guide RNAs targeting those genes and introducing the CRISPR system into hundreds of fish embryos, they identified some animals that developed heart defects as they matured. The team was able to use the DNA barcodes of these fish to track defects up to 13 different inactivated genes.Because of the similarities between zebrafish and humans gene, This finding may point to a previously unknown aspect of human heart development.
Peterson and Parvez are eager for MIC-Drop to work in other labs, saying that the 188 gene screen is still in its infancy. “Ultimately, people want to be able to do genome-wide screening,” says Peterson. “I think this technology will allow us to actually imagine the scale.”
This study is published as “MIC-Drop: Platform for Large In vivo CRISPR Screens”.
MIC-Drop: Platform for large in vivo CRISPR screens, Chemistry (2021). DOI: 10.1126 / science.abi8870
University of Utah Health Sciences
Quote: A new CRISPR-based technology (August 19, 2021) that accelerates the identification of genes involved in health and illness is https: //phys.org/news/2021-08-crispr-based-technology-identification -Obtained from genes on August 19, 2021-involved.html
This document is subject to copyright. No part may be reproduced without written permission, except for fair transactions for personal investigation or research purposes. The content is provided for informational purposes only.
New CRISPR-based technology that speeds up the identification of genes involved in health and disease
Source link New CRISPR-based technology that speeds up the identification of genes involved in health and disease
Read more here: Source link