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A significant accomplishment with CRISPR has been recently achieved. You may have heard of it before this post even, the news has been blowing up with the topic.
But as usual here on Bioscription, we’ll be going a bit deeper into the actual science of the topic, rather than a cursory discussion. Come along, friends.
Researchers at the Northwest A&F University in China were successfully able to implant a gene modification into the genome of cows and then produce fertilized embryos that developed into cows with the modified genes.
The target gene added in this case was a tuberculosis resistance gene called NRAMP1. They had inserted it using a new modified version of CRISPR named Cas9n. This version was able to add the target gene without any off-target effects on the genome whatsoever, an incredible level of precision that many people working on modifying CRISPR have been striving to achieve.
In addition, they were able to precisely add the gene into the segment of the genome they desired. They used a sequencing method called chromatin immunoprecipitation sequencing (ChIP-seq) to find a genome segment that wouldn’t affect any other gene functions.
Though they also wanted a part of the genome that would be properly expressed. Their concerns were that gene silencing from other cellular factors would inhibit the inserted gene’s expression. To avoid this, they added it into the actin gene region, which has strong expression in many tissues across the body.
The scientists used bovine fetal fibroblast cells (BFFs) as the base scaffolding in order to develop a fetus after the gene insertion. They also showed that all was required to insert the gene was to break a single strand of the genome and insert it at that point. The insertion itself is naturally stimulated from the break.
The researchers produced 11 calves with the gene addition and none of them had any off-target effects after sequencing, in direct contrast to when regular versions of CRISPR/Cas9 are used.
The final step was to see if the new gene worked as expected by exposing the calves to Mycobacterium bovis that causes bovine tuberculosis. In this, their gene selection and disease choice was particularly well thought out.
They wanted a gene that would give an innate immunity without disrupting the rest of the bovine immune system. NRAMP1 accomplishes that, as its very expression diminishes the capability for M. bovis to multiply and spread.
In their testing, they found that the gene indeed eliminated intracellular mycobacteria and reduced the growth rate of the rest of it. The percentage of macrophages that were killed in fights with the mycobacteria were also lower as compared to control group non-transgenic cows (From 7.4% to 4.2%).
While the production of tuberculosis-resistant cows is a great success, the true benefit from this experiment is the creation of a modified CRISPR system that has no off-target effects, dramatically increasing its usefulness.
We should expect to see many more experiments utilizing this version (assuming they loan or sell it out to other scientists) in the future.
Photo CCs: Cow Portrait from Wikimedia Commons