• Biology

  The prospects for gene genetics: gene isolation

  Posted by Encyclios on May 16, 2023 at 1:35 PM

  Some genetic diseases can be associated with major changes in the structure of the chromosomes themselves, such as deletions of fragments large enough to be visible under a microscope, duplications or translocations of entire regions from one chromosome to another. This makes it possible to limit the region containing the genetic disease locus and use markers specific to that chromosome. In this way, T. Kunkel’s group at Harvard cloned the Duchenne muscular dystrophy gene whose locus is located on the X chromosome.

  Some patients with that disease showed a rather large deletion right in the X chromosome, and using genetic engineering techniques, the missing fragment was isolated. Then, by comparing the DNA of these patients with that of other patients, who did not show the deletion, it was possible to isolate the gene responsible for the disease. Cloning the muscular dystrophy gene now makes it possible to diagnose it in individuals at familial risk. The history of the study of Duchenne muscular dystrophy provides us with an example of a new methodology in modern genetics called reverse genetics.

  The deletion of the DNA fragment in the muscular dystrophy gene encodes for a protein later called dystrophin whose function was not known. After isolation of the gene using the techniques described above, it was possible to define the function of dystrophin and to establish the molecular and biochemical causes of the disease. With the sequencing of the entire genome of the yeast Saccharomyces cerevisiae and other organisms and the large amount of data already available for the human genome, reverse genetics represents the methodology to study the functions of unknown genes. Through site-specific recombination techniques, it is possible to inactivate the gene of interest and analyze its effects on the organism.

  The technique, called gene knockout (gene inactivation), is widely used in yeast and mice and is a very powerful tool for revealing the basic mechanisms of cellular processes. In the mouse, gene knockout enables the study of molecular mechanisms of development and behavior. Mutated or inactivated genes are introduced into embryonic stem cells, which are reintroduced into mouse embryos at the earliest stage of development. The resulting mice are called chimeras and will contain normal cells and cells with the mutated gene. These cells will help form both the germ line (sperm and eggs) and the somatic line. The resulting animals are paired together to determine whether the mutation of interest has been introduced into the germ line.

  Animals heterozygous for the mutation are paired together to produce a homozygote of the gene of interest. In addition to studying the functions of genes whose phenotype is unknown, this methodology is a very useful genetic tool for studying inherited diseases and developing new protocols for their treatment. After isolating the gene homologous to that in humans for cystic fibrosis, researchers produced mice homozygous for this mutation. These mice exhibit the same symptoms as the human disease and are an excellent model for studying possible therapies, such as gene therapy.

  Among the more recent branches of genetics is genomics, which deals with the study of the genome of living organisms from the perspective of its structure, content and evolution. Underlying genomics are the methods of gene cloning and DNA sequencing. Particularly relevant is the Human Genome Project, which, initiated in 1986, led between 2001 and 2003 to the complete mapping of the human genome that opened up new perspectives in genetic research.

  Today, the boundary between genetics and related disciplines is as thin as ever and destined to become even thinner. In particular, with molecular biology and biochemistry, the areas of research are actually overlapping. In 2020, the scientific community focused the efforts of numerous players (universities, institutions, foundations, pharmaceutical companies) on genetic studies aimed at finding new diagnostic and therapeutic solutions against the Sars-CoV-2 virus responsible for the Covid-19 pandemic.

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