By IMBA-Institute of Molecular Biotechnology of the Austrian Academy of Sciences
Many basic principles of biology and basically all pathways that regulate development are identified in so-called genetic screening. Gene screening was originally the pioneer of Drosophila fruit flies and the nematode Caenorhabditis elegans, and its gene screening involved the inactivation of many genes. By analyzing the consequences of gene loss, scientists can draw conclusions about its function. In this way, for example, all the genes required to form the brain can be identified.
Genetic screening can usually be performed in flies and worms. In humans, there is a lot of knowledge about genetic diseases and the consequences of disease-related mutations, but systematic analysis is not possible. Now, IMBA̵
In the Jürgen Knoblich group of IMBA, by using brain organoids (a 3-D cell culture model for the human brain), CRISPR-LICHT can now be used to analyze the effects of hundreds of mutations in the human brain.
Co-first author Dr. Dominik Lindenhofer explained: “The basis of the technology is to combine the famous CRISPR-Cas9 technology with the double barcode that won the Nobel Prize in October 2020.” IMBA student. “The key trick is to apply guide RNA and genetic barcodes, which is a type of DNA that we add to the cell genome used to grow organoids. This allows us to see the entire cell lineage of each organoid, and The second barcode allows us to count the number of cells produced by each starting cell, thereby reducing noise, so we can determine the effect of each guide RNA on the number of cells produced during the growth of organoids. Cell resolution in heterogeneous tissues CRIPSR Lineage Tracking (CRISPR-LICHT).”
The researchers applied CRISPR-LICHT to microcephaly, a genetic disease caused by the shrinking brain size of patients and severe mental disorders. Through this revolutionary new technology, scientists have screened out all genes suspected to play a role in the disease.
IMBA postdoctoral and co-first author Christopher Esk said: “We were able to use CRISPR-LICHT not only to identify microcephaly genes, but also to identify specific mechanisms that control brain size.” Endoplasmic reticulum (ER) It is identified as the main hub that controls the secretion of extracellular matrix proteins. This mechanism affects the integrity of tissues, thereby affecting the size of the brain, and is thought to be a cause of microcephaly.
Gene screening in Drosophila has long been an established tool for genome-wide screening and has a long tradition in Vienna. The Vienna Drosophila Research Center (VDRC), jointly developed by IMBA scientists, is the only fruit fly reserve center in Europe and one of the largest fly collections in the world for functional gene research. Jürgen Knoblich, scientific director and team leader of IMBA, is also rooted in Drosophila genetics. Thanks to Drosophila, he has an important understanding of the role of stem cells in brain development.
“We are pleased to report that we can now routinely perform genetic screening in complex organoid systems. This method can be applied to other organoid models and any diseases that affect organ formation. This is an analysis of brain diseases and burdens. The new method has great future potential because it can be applied to any brain disease including autism. Only with the collaborative spirit of the Vienna Biological Center, our work is possible because of our neighboring Max Perutz laboratory and The research team of the Institute of Molecular Pathology, the lead author of the study, said Jürgen Knoblich.
Building a better brain-the organoid upgrade of bioengineering
C. Esk et al. Human tissue screening identified the modulator secreted by ER as a determinant of brain size. science (2020). science.sciencemag.org/cgi/doi…1126 / science.abb5390
Provided by IMBA-Institute of Molecular Biotechnology, Austrian Academy of Sciences
Citation: A breakthrough genetic screening tool for human organoids (October 29, 2020), retrieved from https://phys.org/news/2020-10-groundbreaking-genetic-screening-tool-human.html to 2020 October 30
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