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published on 22 October 2018 in life

Crispr-Cas9, the gene factory

DNA is like an alphabet with only four letters (G, T, A, C) written one after another. A, C, G, T are the initials of the four molecules that encode life: adenine, cytosine, guanine and thymine. Four letters that, combined together, form genes: the words of the encyclopaedia that give inanimate material the instructions to become life. A gene of our genetic code is a little piece of the long DNA chain contained in the nucleus of almost all cells; human red blood cells, for example, have no nucleus. Each gene guides cell organelles to assemble a specific protein or a specific enzyme, that is the building blocks of our bodies and the nano-machinery that make it work. This inner mechanism has worked for four billion years, that is since the first bacterial cells appeared in the warm puddles of an as yet very young Earth.

Sequence of DNA nucleotide bases

The cell organelles that produce proteins and enzymes are immersed in the cytoplasm, that is in the gelatinous matrix located between the nucleus and the membrane delimiting the cell. DNA never leaves the nucleus, so how does it communicate the assembly instructions to the organelles? RNA, which is an exact copy of a gene, does it. RNA is created directly on DNA by means of an enzyme that reads the original as a strand and composes the copy letter after letter. When the RNA is ready, it detaches itself from the nucleus and leaves it; therefore, the original (DNA) remains safely within the nucleus, while the copy (RNA) is sent into the cytoplasm to tell the nano factories how to produce the proteins.

DNA was first isolated by the Swiss biochemist Friedrich Miescher in 1869, but only a century later, in 1953, the American and British scientists James Watson and Francis Crick revealed its structure and how it functions and for this were awarded the Nobel Prize in 1962. The first complete reading of an entire human genetic code was completed in 2003 as part of the Human Genome Project (HGP). From the end of the 1960s, genetic engineering has manipulated DNA to confer new characteristics to cultivated plants and reared animals or to induce bacteria to produce useful molecules such as insulin: not at all similar, however, to the often marvellous, and sometimes dramatic, consequences of DNA manipulation and mutation described in science-fiction films. But then came Crispr-Cas9: genetic editing.

A Danish dairy company developed this technique in 2012, studying antiviral defence mechanisms in the milk enzymes of yoghurt. Even bacteria fall ill and, to defend themselves from viruses, develop Cas9, a protein that recognises, cuts and eliminates the virus’s DNA sequence that parasites the bacteria genome after infection. How does Crispr-Cas9 work? An RNA molecule complementary to the segment of DNA to be cut is artificially constructed in a laboratory.  A Cas9 originating from bacteria is attached to this RNA. It is then injected into the nucleus of the cell and nature is allowed to take its course:  RNA recognises and binds itself to the original segment on the DNA, and Cas9 cuts it. Now it is possible to wait for the repair systems to reconstruct and adjust the piece of DNA or to take advantage of the cut to insert new genes. The Crispr-Cas9 technique is able to repair or eliminate a defective gene, replace a gene with another able to give the organism new powers and mix genes belonging to different plants or to different animals. The fields of application are countless and certainly, for the most part, unexplored. The technique could be used to cure genetic diseases, to improve the characteristics of animals and food crops, to produce new medicines and to fight ancient ailments.

The most dangerous animal in the world is the mosquito. Nature has made it the perfect vehicle for diseases like Zika virus, yellow fever, dengue fever and chikungunya. Each year, malaria alone year kills half a million people. There are no vaccines, so we can only repel the insects using sprays, defend ourselves from stings, drain the damp areas where they reproduce or use preventive medicines to repel plasmodium, the micro-organism that causes the disease. Mass control with traditional methods is very complicated, and for this reason research is experimenting with  genetic editing techniques to make the mosquitoes unable to transport plasmodium. The idea is to release resistant mosquitoes into the environment so that, when they mate with the wild populations, they will spread the mutation turning the mosquitoes into harmless and only irritating insects. CRISPR technology is also economic and easy to apply. This is certainly a further advantage, although some fear that ill-intentioned and amateur biotechnologists may play around with genes in their kitchens and produce new ailments, invincible bacteria, killer mosquitoes or mutant creatures. But this, fortunately, is still science fiction.

by Andrea Bellati

 
With the sponsorship of the Italian Ministry of Education, Universities and Research
 
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