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Transgenic animals in biomedicine and agriculture, the future and challenge for the farm industry

时间:2011-07-25 14:32:56  来源:  作者:邹贤刚
Xiangang Zou  
The Babraham Institute, Cambridge, CB1 3QZ
Zuohua Liu
Chongqing Animal Husbandry Science Institute
 
1. Introduction: traditional breeding and biotechnology
       Traditional animal Breeding has a long standing and very successful achievements. It began with domestication by which man habituated animals to live in his proximity. The selection was predominantly done on the basis of the phenotype and specific traits. But a scientifically based animal breeding has existed for approximately 50 years on the basis of the increasing knowledge in population genetics and statistics. The biotechnology in farm animal comprises artificial insemination (AI), embryo transfer (ET), cryopreservation, sexing, in vitro fertilization (IVF), embryo bisection, nuclear transfer (NT, cloning) molecular biology and transgenic technology. There are several methodologies that can be used for the production of transgenic animals, including: (1) DNA transfer by retroviruses, (2) microinjection of genes into pronuclei of fertilized zygotes; (3) injection of embryonic stem (ES) cells and/or embryonic germ (EG) cells, previously transfected with foreign DNA, into the cavity of blastocysts; (4) sperm-mediated exogenous DNA transfer during in vitro fertilization; (5) liposome-mediated DNA transfer into cells and embryos; (6) electroporation of DNA into sperm or embryos; and (7) nuclear transfer (NT) with somatic cells, ES or EG cells.
2. Recent strategies for producing transgenic farm animals
     The early method employed widely to produce transgenic animals is called microinjection, which is to purify the gene fragments from a mammalian expression vector and injected the DNA into the male pronuclei of the zygotes, the injected embryos were then transferred into recipients in estrous synchronization. After full term pregnancy, the pups were born and DNA samples were collected from the individuals and were analysed for their transgenic status. The transgenic efficiency for the farm animals, especially for cattle, sheep, goat and swine (there are dark organelles in their zygote of those animals, which block the view of pronuclei under microscope during microinjection) is very low (about 1-10%) and it is also not economical. Two recent advances in somatic nuclear transfer (cloning) and stem (including embryonic and germ) cell biology have profoundly impact on the production of transgenic technology in farm animal. Briefly, DNA can be first transferred into in vitro cultured cells and positive transgenic colonies were selected in vitro and then those cells were used for nuclear transfer procedure to produce transgenic animals. The advantages of the new procedures allow scientists to add gene(s) into in vitro cultured cellular genomes at random or at a specific locus (gene target), which result the production of transgenic animal at 100% efficiency and a gene target farm animal could be produced for the first time by this method.
3. Current progress of transgenic farm animals
       Scientists in Australian have generated transgenic pigs containing an hMt-pGH construct which can tightly be regulated by zinc feeding. The transgenic animals show significant improvements in growth rate, feed conversion and bodyfat–muscle ratio. Transgenic sheep carrying a keratin-IGF-I construct show expression in the skin and the clear fleece was about 6.2% greater in transgenic versus non-transgenic animals. There were no adverse effects of the transgene on health or reproduction were observed. Another interesting application in transgenic farm animals could be enhanced their disease resistance. A mouse model, in which recombinant monoclonal antibodies, which neutralize the transmissable gastroenteritis virus (TGV), are secreted into milk, provided passive protection against gastroenteric infections to the pups. The verification of this model in pigs is promising. Recently, transgenic pigs expressing a bacterial phytase gene under the transcriptional control of a salivary gland-specific promoter were shown to have improved phosphate uptake. The inserted phytase gene was almost exclusively expressed in the salivary gland and enabled the pigs to digest phosphorus in phytate, which could then be metabolized by the intestine. These animals require significantly fewer inorganic phosphate supplements, release substantially reduced phosphorus levels in manure and thus reduce environmental pollution. Transgenic animals expressed bovine alpha-lactalbumin and human lactotransferrin in the mammary gland of cattle, goats or sows were produced and their lactation performance was improved with respect to milk composition, milk yields and offspring survival and growth.
       Gene pharming is to target the expression of the valuable gene(s) to the mammary gland via the use of mammary gland-specific promoter elements, large amounts of transgenic farm animals, such as cattle, sheep, goat, pig, rabbit and chick, have been produced. Human lactoferrin, anti-thrombin III (ATIII), α-anti-trypsin (α-AT), clotting factor IX, tissue plasminogen activator (tPA), human serum albumin (HSA) and many antibodies are produced in the milk already or their researches are current underway. The advantages for producing human protein in the animal milk are: a large amount of proteins can be produced at a low cost; the protein in milk can be purified easily and the protein produced in the mammary gland can maintain its nature folding and biology activity.
       Transgenic and/or gene target pig can be used in xenotansplantation. Organ transplantation has saved many lives so far. But there is a constant shortage of organs available for transplantation. Many patients died while waiting because of the shortage of organ donation. Scientists have proved that pig is the best animal for organ donation in xenotansplantation, mainly because the porcine organs have a similar size as human organs and its anatomy and physiology are not too different from those in humans. For xenotansplantation from pig to human, many studies are focused on hyperacute rejection response (which occurs within seconds or minutes); acute vascular rejection (within days); cellular rejection (within weeks) and chronic rejection (several years). Other researches on prevention of transmission of porcine endogenous retroviruses (PERV) to the human recipient are also underway. In order to over come those difficulties, transgenic and gene target pig models are proposed. Transgenic pig expressing human decay accelerating factor (DAF), human CD46 and CD59 were produced separately, and the survival rate of the transgenic porcine organs after transplantation to primate recipients were increased. For example, transplantation of hDAF-transgenic porcine kidneys was compatible with an extended survival of the recipients, the physiological function of the kidneys was maintained in the primate for up to 3–4 weeks. Another promising strategy towards successful xenotransplantation is the knockout of the antigenic structures on the surface of the porcine organ, 1,3-α-galactosyltransferase. Recently, the generation of piglets by gene target in which both alleles of the α-galactosyltransferase locus had been knocked out, were reported. The usefulness of organs from these pigs for xenotransplantation is currently being tested.
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