Simple about the difficult: how do I edit a gene using CRISPR cas??

 In short, CRISPR — or rather, Crispr cas 9 — is a powerful genome editing tool. It is based on an element of the bacterial defense system that biologists have adapted to make changes to the DNA of plants, animals, and even humans. The technology allows you to make corrections in just a few days, not weeks or months. Never before has humanity had such a precise tool for manipulating genes.

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The history of CRISPR began in 1987, when Japanese scientists studying Escherichia coli found unusual repeating sequences in its DNA. Their biological significance could not be determined, but soon similar fragments were found in the genome of other bacteria and archaea. The sequences are called CRISPR-short palindromic repeats, regularly arranged in groups (Clustered Regular Interspaced Short Palindromic Repeats).

Their function remained a mystery until 2007, when experts on the bacterium Streptococcus, which is used to make fermented milk products, determined that these fragments are part of the bacteria's immune system.

The fact is that bacteria must constantly repel the attacks of viruses — their natural enemies. To do this, they produce special enzymes. Each time a bacterium manages to kill a virus, it cuts up the remnants of its genetic material and stores them inside CRISPR sequences. This information is then used in the event of a new virus attack. When attacked, the bacterium produces Cas9 proteins that carry a fragment of the virus's genetic material. If this site and the attacking virus's DNA match, Cas9 cuts the latter's genetic material and neutralizes the threat.

For a while, this discovery was interesting only to microbiologists. However, everything changed in 2011, when biologists Jennifer Doudna and Emmanuel Charpentier decided to study the mechanism of CRISPR more precisely. They found that the Cas9 protein can be tricked by giving it artificial RNA. A protein carrying such RNA will look for genetic fragments that match what it carries. When it finds a match with someone else's DNA, it will start shredding it, regardless of whether it belongs to a virus, plant, or animal. As noted in a 2012 paper by Doudna and Charpentier, this mechanism can be used to cut any genome in the right place.

In February 2013, it was proved that CRISPR/Cas9 can be used to edit DNA in mouse and human cell cultures. Moreover. it turned out that the technology allows not only to remove unnecessary genes, but also to insert others in their place. To do this, it is enough to add enzymes that restore DNA.

Scientists quickly realized the huge promise of CRISPR. If in 2011 only 100 works about it were published, by 2017 this figure reached more than 14,000.

Among other things, they described analogs of Cas9 proteins. For example, Cas13 can edit RNA rather than DNA.

The idea of gene modification is not new, and its various methods have been around for many years. However, CRISPR surpasses all known technologies so far due to its availability and accuracy. Editing a single gene will cost only $75 and take several hours. And, importantly, the technology works with any organism on Earth.

There are almost an infinite number of potential applications of the technology. First, CRISPR allows scientists to figure out the function of different genes. It is enough to simply cut the studied gene from the DNA and see what body functions were affected. However, the public is much more interested in practical applications. They can be divided into several points:

1) Changes in agriculture

CRISPR allows you to make crops more nutritious, more delicious and resistant to heat and stress. You can also give plants other properties: for example, cut out the allergen gene from peanuts, and introduce resistance to a deadly fungus in bananas. The technology can also be used to edit the genome of domestic animals — such as cows.

2) fight against hereditary diseases

Scientists intend to use CRISPR to cut out mutations from the human genome that are responsible for a number of diseases, such as sickle cell anemia. The technology will also allow cutting out genes for Huntington's chorea or BRCA-1 and 2 mutations associated with breast and ovarian cancer. In theory, a CRISPR attack can even stop the development of HIV.

However, experts will have to make sure that the technique is safe before it is used in hospitals. Cas9 enzymes sometimes make mistakes and edit DNA in unexpected places, which can cause serious health consequences.

3) New antibiotics and antivirals

Bacteria develop resistance to antibiotics, and developing new ones is expensive and difficult. CRISPR technology makes it possible to destroy certain types of bacteria with high accuracy, although a specific technique has yet to be developed. A number of researchers are also working on crispr systems that target viruses.

4) Genetic drive

Using CRISPR, you can change not just the genome of an individual animal or plant, but also the gene pool of an entire species. This concept is known as"genetic drive". Usually, any organism passes on half of its genes to its offspring. However, using CRISPR can increase the probability of gene transfer by inheritance to almost 100%. This will allow the desired trait to quickly spread throughout the population.

Using this technology, you can, for example, modify mosquitoes so that only females are born in their population. After some time, the population will disappear.

In a more gentle version, you can make mosquitoes resistant to infection with malarial Plasmodium. They will not be able to transmit the parasite to humans, and malaria will end. However, to implement such projects, it is necessary to overcome the doubts of skeptics who protest against such a large-scale invasion of nature.

5) the Creation of "designer babies»

This item attracts the most public attention. However, according to scientists, so far our technological capabilities do not allow us to create children with the specified qualities. For example, thousands of genes are responsible for the level of intelligence, and it is not yet possible to correct them all. Perhaps in the future, the technology will reach the desired level, but for now there is nothing to worry about.

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Not all scientists consider CRISPR a safe technology. For example, according to recent research, gene editing can cause extensive non-targeted mutations. The authors of another paper note that CRISPR is wrong in 15% of cases.

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