Genetically Modified Mice — The Classic Products of Transgenic Technology
About genetically modified mice
In 1974, Rudolf Jaenisch created the first mouse carrying a foreign gene by injecting the DNA of the SV40 virus into the blastocyst of the mouse. Later, researchers injected the Murine leukemia virus into mouse embryos to obtain mice that can be stably inherited through the reproductive system, and the foreign genes can be stably expressed in the offspring. These mice, which are stable in inheritance and express foreign genes, are genetically modified mice that we now generally refer to.
Preparation technique
There are two main types, DNA pronuclear microinjection and embryonic stem cell blastocyst microinjection.
DNA pronuclear microinjection
The foreign gene is directly injected into the fertilized egg by a micromanipulator, and the foreign gene is integrated into the DNA to develop into a transgenic animal. This method is the most classical method for making transgenic animals, and it is also the most widely used method. The research on transgenic animals is mostly based on the improvement of Palmer and other methods. Animal species prepared by this method belong to the category of narrowly defined transgenic animals. The genetically modified mice prepared by this method are usually inserted into the gene of interest in a form of multiple copies of the end-to-end form, and the specific mechanism of integration into the genome has not been fully studied.
Embryonic stem cell blastocyst microinjection
The foreign gene is introduced into the embryonic stem cells in vitro, and then the transgenic embryonic stem cells are injected into the animal blastocyst by a micromanipulator, and the
embryonic stem cells can participate in the embryonic composition of the host to form a chimera until the germline chimerism is reached. Embryonic stem cells remain undifferentiated when cultured in vitro. When injected into the blastocyst, they can participate in the formation of various tissue chimeras including the gonads. Therefore, the transfer of foreign DNA into embryonic stem cells can achieve gene transfer. The reproductive system of a born animal may integrate a foreign gene, and an individual having a homozygous gene, that is, a transgenic animal, can be obtained by cross breeding. Embryonic stem cell-mediated methods are more mature in mice and applied later in large animals. Such methods are commonly used for knockout and capture of genes.
Breeding of mice and screening of homozygotes
Since the advent of genetically modified mice, it has been involved in the breeding of transgenic mice and the screening of homozygotes, especially for the mouse model for commercial application. Compared with non-transgenic animals, transgenic animals have unique genetic characteristics: transgenes are known foreign genes; they are randomly integrated into mouse chromosomes; integration of transgenes is mostly single-site, and a few are multi-site integration. The integrated copy number has a single copy and multiple copies. Therefore, the descendants of the original founder transgenic animals as the same ancestor are all monophyletic.
Therefore, screening of homozygous for genetically modified mice must be performed in a single line. Homozygous genetically modified mice have the following advantages:
(1) Eliminating complicated and expensive detection;
(2) Obtaining individual animals with completely identical genotypes;
(3) Facilitating animal conservation and the like.
However, the following problems are often encountered during the homozygous screening of genetically modified mice:
(1) Screening of homozygous genetically modified mice is a single line transfer, kinship mating, which brings a big contradiction, that is, the transgene can only mate between siblings (full siblings or half siblings) during homozygous process. However, this mating method will increase the inbreeding coefficient of the germline itself;
(2) The foreign gene introduced during the passage may be mutated, resulting in a change in the desired trait.
Application in the field of medicine
With the development of trans-discipline of transgenic and experimental animals, genetically modified mice have been widely used in the field of medicine.
Genetically modified mice and viral diseases
Hepatitis B virus (HBV) infection is a serious threat to human health. The global hepatitis B patients are about 360 million. It is easy to form acute and chronic hepatitis, liver cirrhosis and even liver cancer. The host specificity of the virus has restricted the application of animal models, such as HBV (WHV), duck HBV (DHBV), and human HBV, which can not reflect the replication of HBV in the human body. Therefore, it is urgent to find a small animal model with clear genetic and immune background. Genetically modified mice are undoubtedly a good choice.
Genetically modified mice and tumor research
Cancer in genetically modified mice is transformed from normal cells in a normal microenvironment, so transgenic mouse models can accurately mimic human tumors. Moreover, researchers can monitor and analyze various stages of tumor development, including early tumor development, especially for the analysis of certain landmark activities in tumor development (such as metastasis, angiogenesis, etc.). In addition, genetically modified mice can be used in the development of tumor treatment methods. The transgenic mouse model is an extremely useful model for studying human tumors.
Application of genetically modified mice in pharmacology
In the field of pharmacology, genetically modified mice have two main uses: first, the use of transgenic mice for drug efficacy evaluation; second, the early detection of drug toxicity through toxicological testing. Although the use of animal experiments to predict the efficacy and safety of drugs has been questioned, it is generally believed that pharmacokinetic data can be reasonably extrapolated to humans. In vitro studies, the use of human liver microsomes, human hepatocytes, liver sections and recombinant enzymes are also important methods for assessing human drug metabolism. However, it is impossible to predict how the process of drug absorption, distribution, metabolism and excretion (ADME) in the body affects pharmacological activity and toxicity. The only way to overcome this problem is to study the process of new drugs in intact animals.