In a study published in the June 2011 issue of Science, researchers from University of Washington experimented with fruit flies that could be put to sleep on demand. Flies that underwent 4 hours of induced sleep after training formed long-term memories better than those that were not allowed to sleep after training sessions. Importantly, training alone was not enough to trigger memory consolidation. Sleep was a necessary component.

Many human studies also show that sleep improves memory and performance. Not only may sleep help consolidate memory, but lack of sleep may also hurt it. A 2010 study from Biological Psychiatry found that chronic insomnia may lead to loss of brain volume. Researchers used MRI scans to examine the brains of 37 human subjects with and without chronic insomnia. Insomniacs had smaller volumes of gray matter in three brain areas. The more advanced was the insomnia, the greater was the loss of brain volume in these areas. Preliminary 2012 study from the Washington University School of Medicine found that poor sleep may be linked to brain plaques found in people with Alzheimer’s disease.

Objective:
Explain the processes involved in cloning and producing transgenic animals

Genetic modification
Genetic modification is the change of the genes of a living organism such as a plant or animal using modern techniques of biotechnology. DNA sequencing of entire genomes has been instrumental in the development of new genetically modified animals.

Purpose of genetic modification of animals
The main purposes of using genetic modification of animals are:

improving food-producing animals and agricultural plants
Development of new of plants and animals through natural selection and evolution is a slow process that takes millions of years. Ancient people learned to improve the speed through artificial selection. As the demand for food increases with world’s population growing, genetic modification of animals and plants became the most efficient way of increasing agricultural production. Genetically modified animals and plants developed in a matter of a few decades produce more and better milk, meat, fiber, they became more resistant to diseases and droughts.

development of treatments for human disease
With genetic modification of animals, production of certain important human proteins such as insulin has become a reality. Scientists have seen success with the use of some animal tissues, bones, and joints in humans. Research continues in the field of skin and organ tissue transplantations. Pigs are commonly used in this research because of their similarities to humans. Pigs producing ‘humanized’ skin, for example, can be used as a source for skin restoration in humans. Animals such as mice are genetically modified to help understanding and developing treatments for human and animal diseases.

production of pharmaceuticals
Genetic engineering has allowed production of certain hormones (insulin, human growth hormone, bovine and porcine somatotropin and other). Current research has shown the possibility of genetically modifying sheep or cattle to produce human proteins in their milk.

Benefits
Genetic modification of animals has shown several benefits. These include increased resistance to disease and parasites, increased productivity and improved hardiness to weather factors. Other benefits involve animal products, such as increased yields of meat, eggs, and milk.

Cloning animals
There are two main types of animal cloning: reproductive cloning and cloning through recombinant DNA technology

Reproductive cloning is a common process very well known for creating an exact genetic match of an animal. In 1996, Dolly the sheep was produced by reproductive cloning and was the first animal to be cloned from adult DNA. Since Dolly, advancements have been made in the process of cloning, and several other animals have been cloned. The reproductive cloning process begins with the transfer of genetic material from the nucleus of a donor adult cell to an egg whose nucleus has been removed. The egg is stimulated with chemicals or electrical current to promote cell division. As the cloned embryo divides in a test tube and reaches a suitable developmental stage, it is transplanted into a female host. The female carries the cloned embryo until birth.

Recombinant DNA (rDNA) technology is the process that occurs when fragments of DNA from two different species are joined in vitro to form a single DNA molecule. Usually, the DNA from one species is incorporated in a plasmid that is a circular DNA replicating independently on the bacterial genomic DNA. A plasmid is then amplified inside the bacteria, isolated and inserted into the genome of embryonic stem (ES) cells to be altered, allowing the DNA of two different organisms to be combined in a single cell. Then the cell is injected into an embryo at early stage of development and the embryo is then implanted in foster mother uterus to produce a chimeric transgenic animal.

Transgenic animals
A transgenic animal is an animal that has incorporated a foreign gene into its cells. The animal can pass this transgene (altered gene) on to its offspring. Every cell within the transgenic animal contains a copy of this transgene. Several different methods can be used to produce transgenic animals.

Microinjection is the most common technique used to produce transgenic animals. Injecting DNA into a cell using a fine-diameter glass needle and a microscope constitutes microinjection. During this operation, the chosen gene from the same or a different species is directly microinjected into the ovum. The injected DNA integrates randomly with nuclear DNA and its expression is possible only when the foreign DNA is attached to a suitable promoter DNA sequence. There are many examples where different types of animal cells have been microinjected and successfully transferred.

Once a manipulated fertilized ovum develops to a specific embryonic phase, it is transferred to the oviduct of a recipient female. The embryo will develop just as a normal embryo and is typically carried full term.

Marker gene is a gene that helps easily identify animals which successfully incorporated transgene. With the use of marker genes, it is possible not only to determine whether a transgenic animal has received the desired DNA but also whether the genes in the DNA are being expressed. The genetic markers help identifying the location of incorporated DNA in the host genome. The expression of a marker gene can be visualised with staining selected tissues or whole embryos. Polymerase chain reaction (PCR) followed by gel electrophoresis is a technique of choice due to its speed and sensitivity.

Improving domestic animals
Currently the U.S. Meat Animal Research Center (USMARC) has developed DNA markers to identify all cattle in the United States. This method allows improved animal traceability and verification of disease sources. DNA testing also allows producers to base their management and selection process on genetic potential of identified animals.

DNA testing can determine the presence of certain traits and proteins. A common method used in the beef cattle industry involves the presence of the tenderness or marbling genes. This information allows producers to enhance the production of tender meat and meat with optimal marbling. It also allows producers to produce a consistent product.

Xenotransplantation
Xenotransplantation is the transfer of living cells, tissues, or whole organs from one species to another. People who need a kidney for transplantation often chose to use a pig kidney even if that organ was obtained from a genetically engineered pig.

Summary:
With the new technology of genome sequencing and mapping, genetic modification holds the potential for improvements in livestock. The main purposes for using genetic modification of animals are increased production of food-producing animals, treatments for human disease, and production of pharmaceuticals.

Did you know?
1. Identical twins are clones that occurred naturally.
2. Many animal cloning technologies are used for clinical reproductive procedures.
3. The process of cloning in the lab is a very complex procedure.

Checking what you have learned
1. What are the main purposes of genetic modification in animals?
2. What is a transgenic animal?
3. What are the steps involved in cloning an animal through reproductive cloning?
4. What are the steps involved in producing a transgenic animal?
5. How are marker genes used?

Scientists developed genetically modified silkworms that produce a spider silk protein that incorporates into silk fibers and makes them much stronger than natural silk fibers and as tough as spider web. This approach allows producing spider silk fibers on an industrial scale for a range of biomedical, military and textile applications such as wound dressings, artificial ligaments, tendons, artificial blood vessels, tissue scaffolds, microcapsules and even the body armor.

Despite the great demand of the spider fibers and numerous benefits it can offer, it is impossible to farm spiders because of territorialism and cannibalism between spiders. To get around this problem, recombinant spider silk proteins have been produced in other hosts such as bacteria, yeast, baculovirus, mammalian cells, and transgenic plants and animals. However, all these approaches are expensive, hard to scale up, and lack natural spinning of silk fibers. Silkworms represent the perfect host for spider silk production. Their silk glands naturally produce silk fibers. Gene encoding natural silk protein can be replaced with recombinant spider silk protein that can be targeted to the silk gland using a tissue-specific promoter.

In the attempt to generate transgenic silkworms, scientists created a vector that encodes the synthetic spider silk protein A2S814 and targeted its expression to the silkworm’s silk gland as well as an enhanced GFP (EGFP) tag for visual monitoring of the recombinant protein expression. They injected the vector into eggs and selected transformed larvae using EGFP as a marker of A2S814 expression.

Further analyses of silk extracted from the cocoons confirmed that chimeric silkworm/spidersilk proteins expressed by the transgenic larvae and incorporated into chimeric silk fibers. Importantly, the composite fibers were on average stronger than the parental fibers and more extensible than parental silkworm and native dragline spider silk. “These results demonstrate that the chimeric silkworm/spider silk proteins can significantly improve the mechanical properties of composite silk fibers.

Scientists from the Shanghai University of Neuroscience discovered a gene GluR4 that increases the interactions between neurons in the medial prefrontal cortex and multiplies electric signals created in the area. It appears that the expression level of the gene is associated with social status of individual mice.

In order to confirm the theory, the scientists crated transgenic mice that express high level of GluR4 gene in neurons of these regions of the cortex and compared them with the mice of original strain. In order to determine which of the two mouse lines is higher in the hierarchical ladder, scientists conducted a simple test.They allowed the mice of these two strains to meet in a narrow passage way leading to food and looked what mice could grab more food. The results of the experiments confidently showed that mice with over-expressed GluR4 protein reached the food much faster. After a more detailed analysis of brain tissue, the researchers noticed also that the cells of the medial prefrontal cortex of the mutant mice have a higher ability to transmit the signals.

In addition, the opposite effect was achieved after mice were transfected with the gene R4Ct, which suppresses the transmission of neural signals. These mice became less successful in reaching the food.

If one looks at this finding from human perspective, it becomes clear that our leadership capabilities depend, in part, on the expression level of the gene. Would it be useful to screen people for the level of expression of this gene before offering them jobs that require leadership abilities?