The study find that the the exact consequences caused by the loss or addition of that genetic information. In answer, the realm of genetics has been revolutionized by the emergence of the powerful gene editing tool called CRISPR-Cas9, which allows scientists to insert or remove single genes with ease and precision across a wide range of cells. CRISPR is chiefly used to introduce a mutation into a targeted gene and thus remove its function from a certain kind of cell.
The powerful gene editing
The particular changes that then occur in the cell; called the phenotype, are then studied so that scientists know what exactly that gene did in the living cell. Scientists have now come up with an exciting new method to achieve a precise genotype-phenotype linkage. In an exciting new study; CRISPR-Cas9 use is follow by laser micro dissection and single cell geno typing. This protocol makes it possible to find out exactly what phenotypic changes the CRISPR-induced mutations caused in the cell studied, and then to confirm the exact nature of the underlying DNA change that produced that phenotype.
This will make CRISPR still more of a dream tool for gene study; especially in the nervous system. CRISPR-Cas9 does its job by cutting the DNA strand in the cell at the target site; following which the cell repairs the strand using a process called non homologous end joining (NHEJ). The problem is that f is not a very precise process; and can itself introduce new material into the DNA strand, or leave some material out; or substitute one small piece of DNA for another. These are all different types of additional mutations that the researchers do not want in that cell.
The nervous system
Another challenge is that CRISPR doesn’t always achieve a complete knockout of the gene targeted in both copies of the gene. If even one copy is still functioning, the phenotype-genotype match becomes difficult. In short; unknown mutations make it difficult to tell if the phenotype observed is due to the known mutation they introduced via CRISPR or the inadvertent changes caused by NHEJ or CRISPR inefficiency itself.
So far there was no way to determine the genotype of single cells that had undergone CRISPR-Cas9 transfection. Immuno histochemistry is widely use to confirm that the target gene has been delete; but it depends on whether the right antibodies are available for the specific proteins encoded by the targeted genes. Even then, the antibodies may not distinguish defective nonfunctional proteins from the intact kind. This makes it difficult to verify whether knockout of the gene truly occurred or not.
The specific proteins
The chief importance of the new experimental pipeline is to study the effect of genes which do not produce overt effects or which are not easily observe. This is often the case in the brain neurons. First author Andre Steinecke says, “Some, especially genes in the brain, don’t have strikingly obvious effects or are very difficult to visualize. Our goal was to create a widely applicable strategy, capable of reliably determining the exact genetic cause and correlate it to observed phenotype.”