PRODUCTION & APPLICATION OF TRANSGENIC ANIMAL FOR IMPROVING LIVESTOCK PRODUCTIVITY & HUMAN WELFARE

Nowadays, breakthroughs in molecular biology are happening at an unprecedented rate. One of them is the ability to engineer transgenic animals, i.e., animals that carry genes from other species. The technology has already produced transgenic animals such as mice, rats, rabbits, pigs, sheep, and cows. Although there are many ethical issues surrounding transgenesis. The emergence of transgenic technology has widened the scope of development in case of farm animals and the advent of new molecular biology techniques has paved way by giving a new dimension to animal breeding. The transgenic technology is one of an important tool to meet the future challenges for increased animal’s production. The biological products from animal source should be handled with safety as they are subject to contamination and could be damaged very easily. Thus, safety guidelines should be developed for the commercial exploitation of recombinant proteins and ensure that the transmission of pathogens from animals to human beings is prevented. Therefore, the genetically engineered animals and biotechnology will play a vital role in the production of pharmaceutical proteins and bring about a complete refinement in agriculture production by increasing the quality and quantity of production, protection of environment, maintenance of genetic diversity and overall improvement in animals welfare. The animal which are genetically engineered, called as transgenic animals, produce and secrete such novel therapeutic proteins throughout their life. This not only enables several human diseases to be cured, but also make many people needing organ transplantation to live normal life. The transgenic technology plays a potential role in livestock improvement programmes and adds a new dimension to animal breeding.

What is a transgenic animal?————-

There are various definitions for the term transgenic animal. The Federation of European Laboratory Animal Associations defines the term as an animal in which there has been a deliberate modification of its genome, the genetic makeup of an organism responsible for inherited characteristics.

A transgenic animal is one whose genome has been changed to carry genes from other species.

The nucleus of all cells in every living organism contains genes made up of DNA. These genes store information that regulates how our bodies form and function. Genes can be altered artificially, so that some characteristics of an animal are changed. For example, an embryo can have an extra, functioning gene from another source artificially introduced into it, or a gene introduced which can knock out the functioning of another particular gene in the embryo. Animals that have their DNA manipulated in this way are knows as transgenic animals.

The majority of transgenic animals produced so far are mice, the animal that pioneered the technology. The first successful transgenic animal was a mouse. A few years later, it was followed by rabbits, pigs, sheep, and cattle

The two most common reasons are:

Transgenic animals are useful as disease models and producers of substances for human welfare.

• Some transgenic animals are produced for specific economic traits. For example, transgenic cattle were created to produce milk containing particular human proteins, which may help in the treatment of human emphysema.

• Other transgenic animals are produced as disease models (animals genetically manipulated to exhibit disease symptoms so that effective treatment can be studied). For example, Harvard scientists made a major scientific breakthrough when they received a U.S. patent (the company DuPont holds exclusive rights to its use) for a genetically engineered mouse, called OncoMouse® or the Harvard mouse, carrying a gene that promotes the development of various human cancers

• How are transgenic animals produced?————–

Since the discovery of the molecular structure of DNA by Watson and Crick in 1953, molecular biology research has gained momentum. Molecular biology technology combines techniques and expertise from biochemistry, genetics, cell biology, developmental biology, and microbiology.

Scientists can now produce transgenic animals because, since Watson and Crick’s discovery, there have been breakthroughs in The insertion of a foreign gene (transgene) into an animal is successful only if the gene is inherited by offspring.The success rate for transgenesis is very low and successful transgenic animals need to be cloned or mated via—

• recombinant DNA (artificially-produced DNA)

• genetic cloning

• analysis of gene expression (the process by which a gene gives rise to a protein)

• genomic mapping

The underlying principle in the production of transgenic animals is the introduction of a foreign gene or genes into an animal (the inserted genes are called transgenes). The foreign genes “must be transmitted through the germ line, so that every cell, including germ cells, of the animal contain the same modified genetic material.” (Germ cells are cells whose function is to transmit genes to an organism’s offspring.)

To date, there are three basic methods of producing transgenic animals:

• DNA microinjection

• Retrovirus-mediated gene transfer

• Embryonic stem cell-mediated gene transfer

Gene transfer by microinjection is the predominant method used to produce transgenic farm animals. Since the insertion of DNA results in a random process, transgenic animals are mated to ensure that their offspring acquire the desired transgene. However, the success rate of producing transgenic animals individually by these methods is very low and it may be more efficient to use cloning techniques to increase their numbers. For example, gene transfer studies revealed that only 0.6% of transgenic pigs were born with a desired gene after 7,000 eggs were injected with a specific transgene

BENEFITS AND RISKS OF TRANSGENIC ANIMALS————

BENEFITS RISKS

Desired characteristic may be introduced for animal that require few feed supplements as well as medical treatments. Insertion of foreign gene may upset the expression of the genome.

A desired characteristic of offspring could be established in one generation. Normal reproduction may result in a transgene being released to the wild population.

The characteristic required can be chosen with greater specificity and accuracy.

Methods for Producing Transgenic Animals: ———-

The main principle in the production of transgenic animals is the introduction of a foreign gene or genes into an animal (the inserted genes are called transgenes). The foreign genes must be transmitted through the germ line, so that every cell, including germ cells, of the animal contains the same modified genetic material . The first transgenic animals produced in 1980, were mice . There are various methods for producing transgenic animals which are summarized in table 2.

TABLE 2: VARIOUS METHODS FOR PRODUCING TRANSGENIC ANIMALS BY INTRODUCTION OF FOREIGN DNA INTO THE MAMMALIAN GENOME

TECHNIQUES REMARKS

Pronuclear micro- injection (introduction of expressed gene)

Technical simplicity; low success rate; applicable to a wide range of species; most widely used; unpredictable effects due to random transgene integration

Embryonic stem (ES) cell manipulation (introduction of expressed gene, or gene inactivation by homologous recombination)

Substitution of a functional gene with an inactive gene; germ-line competent ES cells have been isolated in mice; ES-like cells identified in other species, including primates, but totipotency remains to be established

Cre-lox technique

Preferred method with more control over resulting phenotype; time-consuming

Viral vectors Complex; largely restricted to avian species

Cytoplasmic injection Less efficient than direct pronuclear microinjection

Primordial germ cells Chimaeric animals result

Nuclear transfer Large potential for genetically modifying livestock

Spermatogonial manipulation Transplantation into recipient testes

How do transgenic animals contribute to human welfare?———–

The benefits of these animals to human welfare can be grouped into areas:

• Agriculture

• Medicine

• Industry

Applications of Transgenic Animals:-————–

Livestock Production

There are many potential applications of transgenic technology in producing new varieties of livestock that has increased growth rate, reproductive performance, feed utilization, improved milk production and high disease resistance. Other by products like meat and eggs also could be modified by this technology.

Carcass Composition and Growth Enhancement—-

Transgenic animals with exogenous gene constructs have been produced which has enhanced growth rate and improved quality of food. Growth hormone and insulin like growth factors genes have been expressed at different levels in transgenic animals . Transgenic cattle and salmon fish have been produced that contains foreign gene constructs. The introduction of chicken ski gene has caused muscular hypertrophy in case of pigs and cattle. The acid meat gene or Rendement Napole gene has been involved in low processing yields of pork there by affecting the quality of meat in pig. Silencing the expression of this gene in case of pigs alter the post mortem pH and improve the quality of meat. Other genes like GH releasing factor, IGF binding proteins also play a major role in the modification of growth. Transgenic pig with human metallothionein promoter had a significant improvement in growth rate and feed conversion

Milk production and Lactation———-

The advances in transgenic technology provide ample chances to improve both the quality and quantity of milk produced. The animals could be made to secrete nutraceuticals in milk that may have an impact over the growth of offspring. Casein variants are the main target for improving the milk composition, which in turn alters the physio-chemical properties of milk. Brophy et al. (2003) reported that cloned transgenic cattle have been developed that produce increased amounts of beta and kappa casein in milk that increase the value of milk in the production of milk based products like cheese, yoghurt and also increase the shelf life of milk products. Transgenic animals also could be developed to produce “infant milk” that has increased levels of human lactoferrin, to generate lactose free milk for lactose in tolerance populations by inhibiting the expression of lactalbumin locus and to produce hypoallergenic milk by knocking of down the expression of B-lactoglobulin gene. Transgenic animals could also be made to secrete antibodies in their milk that give resistance against several diseases like mastitis or to secrete antimicrobial peptides like lysozyme. Grosvenor et al. ( reported that the milk composition could also be altered by making the transgenic animals to secrete growth factors in milk, which in turn affect the growth and maturation of newborn offspring.

By-Products –——–

Wool and leather are the major byproducts that could be altered by genetic engineering. The quality, colour, length, ease of harvest and yield of wool have been the target for manipulating the fiber and hair of farm animals. Transgenic sheep expressing keratin – IGF 1 construct have been produced that have greater percentage of clean fleece than non transgenic animals (Damark et al., 1996). The quality of wool could also be increased by transgenic modification of cystein pathway. Farm animals also could be genetically engineered by altering the collagen gene so that the amount of connective tissue within the skin may be significantly changed so as to improve the quality of hide. Hollis et al. (1983) reported that epidermal growth factor gene (EGF) has been introduced in sheep that enhances the ease of removing the wool from sheep.

Disease Resistance———

The most important application of transgenic technology is the manipulation of MHC (Major Histocompatability Complex) genes, which influence the immune response and increase the disease resistance capacity of livestock. Clements et al. (1994) reported that transgenic sheep have been developed that is resistance to Visna virus infection. The transmission of bovine spongiform encephalopathy (Scrapie) is also prevented by the knock down of prion protein. Transgenic mice have been developed that secretes recombinant antibodies in milk to neutralize the corona virus responsible for transmissible gastro enteritis (TGEV), an economically important disease in case of pigs

Reproductive Performance-———–

Several candidate genes have been identified that increase the reproductive performance of farm animals. Rothschild et al. (1994) reported that the polymorphism in estrogen receptor gene (ESR) increased the litter size in case of pigs. The ovulation rate of different breeds of sheep, which are superior in carcass traits and wool production, could be increased by incorporating a single major autosomal gene called booroola fecundity gene (FECB) that increases the prolificacy in sheep (Piper et al., 1985). The estrus symptoms in case of pigs could be enhanced by incorporating a gene from baboons, which make their posterior red

Historically, mice have been used to model human disease because of their physiological, anatomical and genomic similarities to humans. Transgenic animals are produced as disease models (animals genetically manipulated to exhibit disease symptoms so that effective treatment can be studied) such as Alzheimers, cancer, AIDS. Transgenic animals enable scientists to understand the role of genes in specific diseases. The benefits of using transgenic animals include the possibility of the replacement of higher species by lower species- through development of disease models in mice rather than in dogs or non-human primates, the extent of discomfort experienced by parent animals during the experimental procedures. Transgenic animals such as mice have been found to be valuable in investifations into gene function and for analysis of different hereditary diseases

The FDA suggested that cloned animals and their products were safe to eat for human being . Some drawbacks are associated due to their muscle hypertrophy like difficulties in calving requiring Caesareans, poor viability of calves and poor fertility.

Transgenic animals are used for production of proteins such as alpha-1-antitrypsin, produced in liver, used in treatment of emphysema or cystic fibrosis. This process is less expensive than production of protein through culture of human cells . The human lungs are constantly get affected by foreign particles such as dust, spores and bacteria. To prevent these, neutrophils releasing the elastase enzyme but this enzyme harmed the elastin in the lungs which maintains the elasticity of lungs. So, human body releases a protein α1 proteinase inhibitor which has been successfully expressed in sheep . Recombinant human proteins produced in the mammary glands of transgenic animals Pharmaceutical proteins are now used for commercial purpose . Two scientists at Nexia Biotechnologies in Canada spliced spider genes into the cells of lactating goats. The goats are used to manufacture silk, milk and secrete tiny strands from their body by the bucketful. By extracting polymer strands from the milk and weaving them into thread which is light and tough material that could be used to prepare military uniforms, medical micro sutures and tennis racket strings. Americans are more supportive (60%) for above use of transgenic animals. The mammary gland of transgenic goats is used to produce Monoclonal Antibodies. A recombinant bispecific antibody is produced by using transgenic cattles with in their blood

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Another application includes newly generation of trans-chromosomal animals in which a human artificial chromosome containing the complete sequences of the human immunoglobulin heavy and light chain loci was introduced into bovine fibroblasts, which were then used in nuclear transfer. Transchromosomal bovine offspring were obtained that expressed human immunoglobulin in their blood. This could be a significant step forward in the generation of human therapeutic polyclonal antibodies

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Scientist developed the mice by altering the genes of the mousepox virus in Australia . Some scientist also thought to develop genetically modify mosquitoes so they cannot produce malaria but other scientist worry about these mosquitoes that they could have unforeseen possibly risk if, they are released into the environment

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Now a day approximately about 250000 people are alive due to the successful transplantation of an appropriate allotransplantation. Sometimes there is limitation of appropriate organs or rejection of live organ donation. So, to rectify this problem porcine xenografts from domesticated pigs are considered to be the best choice . Pigs which are genetically modified can be used as a source animal for tissues and organs in human beings for transplantation purpose by delete the gene responsible for the human rapid immune rejection response . In Canada, a National survey on xenotransplantation showed that only 48% found acceptable for ‘the use of animals as a source of living cells, tissues or organs to prolong human life . To overcome the Hyperacute rejection & acute vascular rejection, synthesis of human regulators of complement activity are produced in transgenic pigs . Survival rates, after the transplantation of porcine hearts or kidneys expressing transgenic regulators of complement activity proteins to immunosuppressed nonhuman primates, reached near about 23 to 135 days. So, the Hyperacute rejection can be overcome in a clinically acceptable manner . For long term graft tolerance induction of permanent chimerism via intraportal injection of embryonic stem (ES) cells or the co-transplantation of vascularised thymic tissue

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Blood replacement-————

Transgenic swine are used to produce human haemoglobin. The protein obtained from transgenesis could be purified by using procine blood which is similar to human haemoglobin

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Transgenic animals are used in toxicity testing.

Transgenic animals are used for vaccine testing.

What are the ethical concerns surrounding transgenesis?—————-

This article focuses on the benefits of the technology; however, thoughtful ethical decision-making cannot be ignored by the biotechnology industry, scientists, policy-makers, and the public. These ethical issues, better served in their own article, include questions such as:

Ethical concerns must be addressed as the technology grows, including the issue of lab animal welfare.

• Should there be universal protocols for transgenesis?

• Should such protocols demand that only the most promising research be permitted?

• Is human welfare the only consideration? What about the welfare of other life forms?

• Should scientists focus on in vitro (cultured in a lab) transgenic methods rather than, or before, using live animals to alleviate animal suffering?

• Will transgenic animals radically change the direction of evolution, which may result in drastic consequences for nature and humans alike?

• Should patents be allowed on transgenic animals, which may hamper the free exchange of scientific research?

Limitations of transgenices ———

The transgenic technology even though has tremendous applications in livestock improvement programmes, still it has lots of limitations: Insertional mutations resulting in• alteration of important biological processes. Unregulated gene expression resulting in• improper expression of gene products. Possibility of side effects in transgenic• animals like arthritis, dermatitis and cancer etc. Integration of exogenous DNA sequence• in Y chromosome resulting in transmission only to males.

Transgenic technology holds great potential in agriculture, medicine, and industry.

Interestingly, the creation of transgenic animals has resulted in a shift in the use of laboratory animals — from the use of higher-order species such as dogs to lower-order species such as mice — and has decreased the number of animals used in such experimentation,especially in the development of disease models. This is certainly a good turn of events since transgenic technology holds great potential in many fields, including agriculture, medicine, and industry.

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