1. At the molecular level, cloning generally refers to DNA cloning (also called molecular cloning). The meaning is that a particular DNA fragment is inserted into a vector (e.g. plasmid, virus, etc.) by recombinant DNA technology, and then a large number of identical "populations" of the DNA fragment are obtained by self-replication in the host cell.
2. At the cellular level, a clone is essentially a population of cells formed by the division of a single ****same ancestor cell. Each of these cells is genetically identical. For example, a cell clone is a collective of cells with identical genetic backgrounds formed by dividing a cell in culture in vitro for a number of generations. Another example is that in vertebrates, when an exogenous agent (e.g., a bacterium or a virus) invades the body, specific recognizing antibodies are produced by an immune response. All of the plasma cells that produce a particular antibody divide from a single B cell, and such a population of plasma cells is also a cell clone. Cell cloning is a low-level form of reproduction- asexual reproduction, in which the offspring have the same heredity as the parent without the union of the sexes. The lower the level of biological evolution, the more likely it is that this type of reproduction will be practiced.
3. At the individual level, a clone is a group of two or more individuals with identical genotypes. For example, two identical twins are a clone! Because they come from the same egg cell, they have exactly the same genetic background. By this definition, Dolly is not a clone! Because Dolly is just one person. Only if the British embryologists were able to transplant more than two identical nuclei into more than two identical enucleated oocytes, resulting in more than two Dollys with identical genetic backgrounds, could the word "clone" be used. That's why the authors of the sensational paper, published in the February 1997 issue of Nature, did not describe Dolly as a clone.
Also, cloning can be used as a verb, meaning the process of acquiring the DNA, cells, or groups of individuals described above.
II. Cloning Technology
1. DNA Cloning
Now there are many different methods of DNA cloning, and the basic process is shown in the following diagram (not to scale)
It can be seen that the DNA obtained in this way can be used in many aspects of biological research, including the analysis of the base order of the specific DNA and processing, as well as the biotechnology industry, the mass production of valuable proteins and so on. the mass production of valuable proteins in the biotechnology industry, and so on.
2. Cloning of individual organisms
(1) Cloning of individual plants
In the 1950s, botanists used carrots as a model to study the loss of genetic material in differentiated plant cells, and they were amazed to find that a complete plant could be developed from a single highly differentiated carrot cell
! plant! From this, they concluded that plant cells have totipotency. The carrot population that developed from more than two somatic cells from a single carrot had exactly the same genetic background and was therefore a clone. The cloning process of such a plant is a completely asexual reproduction process!
(2) cloning of individual animals
①"Dolly" birth
February 27, 1997, Edinburgh, England, Roslin (Roslin) Institute of Ian Wilmot scientific research group to the world to announce that the world's first cloned sheep "Dolly" (Dolly) was born, the news immediately stirred the world.
"Dolly" is associated with three ewes. One was a Finnish Dorset ewe in her third month of pregnancy and two were Scottish Blackface ewes. Finnish Dorset ewes provide a full set of genetic information, that is, to provide the nucleus (called the donor); a Scottish black-faced ewes to provide a nucleus-free egg cell; the other Scottish black-faced ewes to provide the environment for the development of the sheep embryo - the uterus, is the "Dolly" sheep "Dolly" sheep. The other female Scottish black-faced sheep provides the environment for the development of sheep embryos - the uterus, which is the "birth" mother of the "Dolly" sheep. The whole cloning process is briefly described as follows:
From the mammary glands of the Finnish Dorset ewes to take out mammary cells, and put them into a low concentration of nutrient medium, the cells gradually stopped dividing, this cell is called the donor cell; to a Scottish black-faced ewe injected with gonadotropin, prompting it to ovulation, take out the unfertilized egg cells, and immediately remove the nucleus, leaving a nucleusless egg cells, this cell is called the recipient cell; this cell is called the recipient cell. This cell is called the receptor cell; using the method of electric pulse, the donor cell and the receptor cell are fused, and finally the fusion cell is formed, as the electric pulse can also produce a series of reactions similar to those in the natural fertilization process, so that the fusion cell can also undergo cell division and differentiation like a fertilized egg, thus forming the embryonic cell; transferring the embryonic cell to the uterus of another Scottish black-faced ewe sheep. The embryonic cells were then transferred to the uterus of another Scottish black-faced ewe, where they further differentiated and developed to form a lamb. The resulting "Dolly" lamb has the exact same appearance as the Dorset ewe.
A year later, another group of scientists reported that transplanting the nucleus of a mouse oocyte (a highly differentiated cell that surrounds the periphery of the oocyte) into an oocyte with the nucleus removed resulted in more than 20 fully developed mice. If Dolly is not enough to be called a cloned sheep because there is only one, these mice
are truly cloned mice.
②Basic process of cloning mice by cell nuclear transplantation
In this experiment, the ovarian thalamus cells were obtained through the following process: the female mice were induced into a state of high egg production by several consecutive injections of chorionic gonadotropin. The complex of oocytes and oocytes was then collected from the oviducts of female rats. The oocytes were dispersed by hyaluronic acid treatment. Oviductal cells with a diameter of 10-12 micrometers were selected to be used as nucleus donors (preliminary experiments showed that oocytes that underwent nucleus transplantation rarely developed to the 8-cell stage if nuclei from oviductal cells with smaller or larger diameters were used). The selected oocytes were kept in a certain solution environment and the nuclei were transplanted within 3 hours (in contrast to the mammary cells used as nucleus donors in the acquisition of "Dolly", which were first passaged 3-6 times in culture)
Oocytes (generally at meiosis II) were collected from female mice of different species by a method similar to that described above. The nuclei of the oocytes were carefully removed under the microscope with a fine tube of about 7 micrometers in diameter, trying not to remove the cytoplasm. The nuclei of the oocytes were similarly carefully removed, also trying to remove as much cytoplasm as possible (by making the removed nuclei move back and forth in the glass tube several times to remove the small amount of cytoplasm that was carried). Within 5 minutes after the nuclei were removed, they were injected directly into the oocytes from which the nuclei had been removed. Nucleus-transplanted oocytes were placed in a special solution for 1-6 hours, and then divalent strontium ions (Sr2+) and cytokinin repressor B were added, the former to activate the oocytes and the latter to inhibit polar body formation and chromosome exclusion. The treated oocytes were then removed and placed in a special solution without strontium and cytokinin inhibitor B to allow the cells to divide and form an embryo.
Embryos at different stages (from the 2-cell stage to the blastocyst stage) were implanted separately into the oviducts or uteri of pseudo-pregnant females that had mated with already ligated males a few days earlier to develop. Fully developed fetal rats were surgically removed after approximately 19 days.
Animals currently cloned by embryonic cell nuclear transfer include mice, rabbits, goats, sheep, pigs, cows and monkeys. In China, all the cloned animals except monkeys are available, and it is also possible to clone goats by successive nuclear transfer, a technique that goes further than embryo splitting and will result in the cloning of more animals. Because the more times an embryo is split, the fewer cells per part, the less capable the individual that develops. There is only one animal that has been cloned by somatic cell nuclear transfer, and that is the "Dolly" sheep.
Three, the gospel of cloning technology
1. Cloning technology and genetic breeding
In agriculture, people use "cloning" technology to cultivate a large number of drought-resistant, anti-fallout, anti-pest and disease-resistant high-quality and high-yield varieties, which greatly increased food production. In this regard, China has stepped into the forefront of the world's most advanced.
2. Cloning technology and the protection of endangered species
Cloning technology for the protection of species, especially rare and endangered species is a gospel, has great prospects for application. From a biological point of view, this is one of the most valuable aspects of cloning technology.
3. Cloning and medicine
In the modern era, doctors are able to perform transplants on almost all human organs and tissues. But as far as science and technology are concerned, rejection in organ transplantation remains the biggest headache. The cause of rejection is poor compatibility due to tissue mismatch. If the organs of a "cloned human being" are provided to an "original human being" for organ transplantation, there is absolutely no fear of rejection because the two are genetically compatible and the tissues are compatible as well. The question is: Is it humane to use "clones" as organ donors? Is it legal? Is it economically viable?
Cloning technology can also be used to reproduce large numbers of valuable genes, for example, in medicine, it is through "cloning" technology to produce insulin for diabetes, growth hormone to make people with dwarfism grow taller again, and stem deflectors that can fight a variety of viral infections, and so on.