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INTRODUCTION……….1 HISTORY………..……….2 PROCESS OF GENETIC ENGINEERING………….4 APPLICATION…………….9 CONCLUSION……….…16
This is accomplished by isolating and copying the genetic material of interest using molecular cloning methods to generate a DNA sequence containing the required genetic elements for expression. Genetic engineering techniques have been applied in numerous fields including research. and then inserting this construct into the host organism. Other forms of genetic engineering include gene targeting and knocking out specific genes via engineered nucleases such as zinc finger nucleases or engineered homing endonucleases. The most common form of genetic engineering involves the insertion of new genetic material at an unspecified location in the host genome. however traditional breeding methods are typically used for the propagation of recombinant organisms. Medicines such as insulin and human growth hormone are now produced in bacteria. It involves the introduction of foreign DNA or synthetic genes into the organism of interest. is the direct human manipulation of an organism's genome using modern DNA technology. biotechnology. The introduction of new DNA does not require the use of classical genetic methods. and medicine. experimental mice such as the oncomouse and the knockout mouse are being used for research purposes and insect resistant and/or herbicide tolerant crops have been commercialized.INTRODUCTION G enetic engineering. also called genetic modification. Genetically engineered plants and animals capable of producing biotechnology drugs more cheaply than current methods 3 .
a science started form the early 1900’s based on experiments by the Austrian monk. researchers used recombinant DNA to engineer bacteria to produce small quantities of insulin and interferon. The U. During the late 1970’s. and the other was the 1973 discovery by Cohen and Boyer of a recombinant DNA technique by which a section of DNA was cut from the plasmid of an E. coli bacterium and transferred into the DNA of another. a Stanford professor and Nobel laureate. Genetic engineering based on genetics. Genentech’s Humulin. 4 . DNA is identified as the carrier of genetic information by Oswald Avery Colin McLeod and Maclyn McCarty. by Watson and Crick. the first field tests of genetically engineered crops (tobacco and tomato) are conducted in the United States. Later two important key events happened. a form of human insulin produced by bacteria. In 1982. One of the key scientific figures that attempted to highlight the promising aspects of genetic engineering was Joshua Lederberg. Gregor Mendel.S Food and Drug Administration approve the first genetically engineered drug. green genetic engineering was born. In 1944. Genetic material is introduced into cell cultures for the first time ever with the aid of Agrobacterium tumefaciens. In 1980. Committee of the national Academy of Sciences concluded that transferring genes between species of organisms posed no serious environmental hazards. One was the 1953 discovery of the structure of DNA. In 1987.HISTORY T he origins of biotechnology culminated with the birth of genetic engineering.
The People’s Republic of China was the first country to commercialize transgenic plants. making it the first genetically engineered crop commercialized in Europe. Bt Potato was approved safe by the Environmental Protection Agency. In 1994. Craig Venter Institute announced that they had created the first synthetic bacterial genome. the European Union approved tobacco engineered to be resistant to the herbicide bromoxynil. Paraguay and South Africa. named Synthia. introducing a virus-resistant tobacco in 1992. the largest of which by area grown were the USA. 5 . making it the first pesticide producing crop to be approved in the USA. Canada. and added it to a cell containing no DNA. scientists at the J. In 2010. Argentina. China. In 1994 Calgene attained approval to commercially release the Flavr Savr tomato. In 2009 11 transgenic crops were grown commercially in 25 countries. was the world's first synthetic life form. The resulting bacterium. Brazil. India. a tomato engineered to have a longer shelf life.] In 1995.
In animals the majority of genes used are growth hormone genes. Once isolated. Selection 6. This typically involves multiplying the gene using polymerase chain reaction (PCR). If the chosen gene or the donor organism's genome has been well studied it may be present in a genetic library. Constructs 3. Confirmation ISOLATING THE GENE The gene to be inserted into the genetically modified organism must be chosen and isolated. most genes transferred into plants provide protection against insects or tolerance to herbicides. 6 . but no copies of the gene are available.PROCESS OF GENETIC ENGINEERING There are 7 steps in genetic engineering 1. Presently. the gene is inserted into a bacterial plasmid. Isolating the gene 2. Regeneration 7. Gene targeting 4. it can be artificially synthesized. Once chosen the genes must be isolated. Transformation 5. If the DNA sequence is known.
CONSTRUCT The gene to be inserted into the genetically modified organism must be combined with other genetic elements in order for it to work properly. is needed to determine which cells are transformed with the new gene. such as restriction digests. engineered homing endonucleases. In addition to enhancing gene targeting. The promoter region initiates transcription of the gene and can be used to control the location and level of gene expression. which in most cases confers antibiotic resistance to the organism it is expressed in. The constructs are made using recombinant DNA techniques. The frequency of gene targeting can be greatly enhanced with the use of engineered nucleases such as zinc finger nucleases. Other techniques allow new genetic material to be inserted at a specific location in the host genome or generate mutations at desired genomic loci capable of knocking out endogenous genes. The technique of gene targeting uses homologous recombination to target desired changes to a specific endogenous gene. ligations and molecular cloning. This tends to occur at a relatively low frequency in plants and animals and generally requires the use of selectable markers. As well as the gene to be inserted most constructs contain a promoter and terminator region as well as a selectable marker gene. engineered nucleases can also be used to introduce mutations at endogenous genes that generate a gene knockout 7 . GENE TARGATING The most common form of genetic engineering involves inserting new genetic material randomly within the host genome. The selectable marker. while the terminator region ends transcription. or nucleases created from TAL effectors. The gene can also be modified at this stage for better expression or effectiveness.
Some genetic material will enter the cells and transform them. In some cases the electroporated cells will incorporate the DNA into their genome. with a heat shock or an electric shock. Stressing the bacteria for example. 8 . In biolistics particles of gold or tungsten are coated with DNA and then shot into young plant cells or plant embryos.TRANSFORMATION About 1% of bacteria are naturally able to take up foreign DNA but it can also be induced in other bacteria. which contains T-DNA and can be grown in both E. The genes to be inserted are cloned into a binary vector. In plants the DNA is generally inserted using Agrobacterium-mediated recombination or biolistics. Another transformation method for plant and animal cells is electroporation. can make the cell membrane permeable to DNA that may then incorporate into their genome or exist as extrachromosomal DNA. Agrobacterium naturally inserts DNA from a tumor inducing plasmid into any susceptible plant's genome it infects. Electroporation involves subjecting the plant or animal cell to an electric shock. Once the binary vector is constructed the plasmid is transformed into Agrobacterium containing no plasmids and plant cells are infected. DNA is generally inserted into animal cells using microinjection. where it can be injected through the cells nuclear envelope directly into the nucleus or through the use of viral vectors. which can make the cell membrane permeable to plasmid DNA. In Agrobacterium-mediated recombination the plasmid construct must also contain TDNA. This method can be used on plants that are not susceptible to Agrobacterium infection and also allows transformation of plant plastids. The Agrobacterium will then naturally insert the genetic material into the plant cells. Coli and Agrobacterium. The T-DNA region of this plasmid is responsible for insertion of the DNA. causing crown gall disease.
in most cases a selectable marker is used to differentiate transformed from untransformed cells. As bacteria consist of a single cell and reproduce clonally regeneration is not necessary. In plants this is accomplished through the use of tissue culture. 9 . In animals it is necessary to ensure that the inserted DNA is present in the embryonic stem cells. If successful an adult plant is produced that contains the transgene in every cell. Another method of screening involves using a DNA probe that will only stick to the inserted gene. By growing the cells in the presence of an antibiotic or chemical that selects or marks the cells expressing that gene it is possible to separate the transgenic events from the non-transgenic. All offspring from the first generation will be heterozygous for the inserted gene and must be mated together to produce a homozygous animal.SELECTION Not all the organism's cells will be transformed with the new genetic material. If a cell has been successfully transformed with the DNA it will also contain the marker gene. A number of strategies have been developed that can remove the selectable marker from the mature transgenic plant. REGENERATION As often only a single cell is transformed with genetic material the organism must be regrown from that single cell. Each plant species has different requirements for successful regeneration through tissue culture. When the offspring is produced they can be screened for the presence of the gene.
and the expression and localization of the protein product(s) is usually necessary. To examine expression of the trans-gene. immunofluorescence and phenotypic analysis.CONFORMATION The finding that a recombinant organism contains the inserted genes is not usually sufficient to ensure that the genes will be expressed in an appropriate manner in the intended tissues of the recombinant organism. 10 . further analysis frequently uses PCR. quantitative RT-PCR. RNA processing patterns. which serve to determine the chromosomal location and copy number of the inserted gene. To examine the presence of the gene. an extensive analysis of transcription. Southern hybridization. using methods including northern hybridization. the organism's offspring are studied to confirm that the trans-gene and associated phenotype are stably inherited. and DNA sequencing. When appropriate. Western blot.
To do this artificially may require transferring genes as part of an attenuated virus genome or physically inserting the extra DNA into the nucleus of the intended host using a microsyringe. thereby increasing the amount of arable land on the planet. genes are artificially transferred between organisms that could be conventionally bred. or as a coating on gold nanoparticles fired from a gene gun. Genetically modified foods are one such solution. allow plants to manufacture their own pesticides to ward off insects. novel solutions are increasingly in demand. codon usage of the gene and how to deactivate the gene. In the process of transgenesis. With the world population expanding and synthetic pesticides decreasing in effectiveness. In nature this can occur when exogenous DNA penetrates the cell membrane for any reason. The method to introduce new genes into plants requires several important factors such as specific promoter. In the process of cisgenesis. and allow plants to grow under adverse weather conditions or in poor soil. Genetically modified foods can: increase plants' resistance to pesticides and herbicides. The specific 11 . thereby decreasing the need for these pollutant chemicals. which is a form of horizontal gene transfer.APPLICATION GENETICALLY MODIFIED FOOD AND CROPS O ne major application of genetic engineering techniques is in the realm of food production. Genetic modification involves the insertion or deletion of genes. genes from a different species are inserted. increase the yields of many staple crops and thereby ward off starvation in many areas of the world.
The transgenic genes should also be able to be denatured by heat in order for human consumption. a precursor to vitamin A. strain CP4 Corn Resistance to insect pests. as well as significantly increased growth rates 12 . through expression of the insecticidal protein Cry1Ab from Bacillus thuringiensis Canola High laurate levels achieved by inserting the gene for ACP thioesterase from the California bay tree Umbellularia californica Plum Resistance to plum pox virus conferred by insertion of a coat protein (CP) gene from the virus Rice Three genes for the manufacture of beta-carotene. if we want the gene to express only in the rice instead of the leaf than we would only use an endosperm specific promoter. For instance. in the endosperm of the rice prevent its removal (from husks) during milling Tabacco Hepatitis B virus surface antigen (HBsAg) produced in transgenic tobacco induces immune response when injected into mice Maize Fusion protein (F) from Newcastle disease virus (NDV) expressed in corn seeds induces an immune response when fed to chickens Coho salmon A type 1 growth hormone gene injected into fertilized fish eggs results in 6.promoter must pertain to area that we want the gene to express. The codon usage of the gene must also be more optimize for the rice since there are several different codons for each of the 20 amino acid. Examples of GMOs Resulting from Agricultural Biotechnology Genetically Conferred Trait Herbicide tolerance Example Soybean Genetic Change Insect resistance Altered fatty acid composition Virus resistance Vitamin enrichment Vaccines Oral vaccines Faster maturation Glyphosate herbicide (Roundup) tolerance conferred by expression of a glyphosate-tolerant form of the plant enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) isolated from the soil bacterium Agrobacterium tumefaciens. The reason is because we only want our transgenic genes to express only in the rice and not the leaves.2% retention of the vector at one year of age. specifically the European corn borer.
including frogs. Human identical twins are also clones. rather than an embryo. The real key to cloning an adult animal is the ability to reprogram the skin cell nucleus and cause it to begin developing as if it was a newly fertilized egg. but also raised ethical concerns. the first animal cloned in this way. Cloning creates a genetically identical copy of an animal or plant. but she was not the first. Plants are often cloned – taking a cutting produces a clone of the original plant. a fetus. pigs and mice. This was a major scientific achievement. Cloning from adult animals was introduced to the public in 1997 when scientists announced the birth of Dolly.had been cloned before Dolly. A clone is an animal who is genetically identical to its donor "parent". cattle. Dolly was the first mammal to be cloned from an adult cell. and cows . 13 . We now know that this can be achieved using cells derived from a microscopic embryo. or from an adult animal. identical offspring. Cloning requires specialized microsurgery tools and involves five basic steps: Enucleation of the recipient egg Transfer of the donor cell into the recipient egg Fusion of the donor cell to the recipient egg Culturing the resulting cloned embryo in the incubator Transferring the developing embryo into the reproductive tract of a surrogate mother Dolly the sheep may have been the world's most famous clone. There have now been hundreds of clones produced from skin cells taken from adult sheep. Many animals . mice.CLONNING C loning is the production of multiple. goats.
other sheep have been cloned from adult cells. In 2004 a mouse was cloned using a nucleus from an olfactory neuron. horses and donkeys. goats and cattle. showing that the donor nucleus can come from a tissue of the body that does not normally divide. rabbits.Since 1996. when Dolly was born. Process of Cloning 14 . pigs. as have mice.
parental testing and criminal investigation. though. which can also be used as the person's identifier. An increasing range of applications makes use of this fact. The 1% that’s different includes several frequently repeating sequences. about 99% of human DNA is identical from one person to the next. Therefore. DNA typing. in DNA fingerprinting. and comparing an entire DNA sequence with another would be hard to do. DNA profiles are encrypted sets of numbers that reflect a person's DNA makeup. It is used in. animals. for example. Plants. 15 . they differ mainly in how the fragments are extracted and how they are converted into a form that can be analyzed for identification. DNA profiling should not be confused with full genome sequencing. DNA sequences are extremely long. There are several techniques for doing so. and even bacteria have unique DNA fingerprints. or genetic fingerprinting) is a technique employed by forensic scientists to assist in the identification of individuals by their respective DNA profiles.DNA FINGERPRINTING D NA profiling (also called DNA testing. the number of repeating sequences in any given position on a chromosome is different for each person. While human DNA fingerprinting has numerous uses in law and forensics—from verifying paternity to identifying murder suspects—this technique also applies to other organisms. fragments of DNA are extracted and a collection is created that is unique for each person. Fortunately.
Process of DNA Fingerprinting Examples : 16 .
the technique can be used to develop drugs. diabetes. and to reproduce important human hormones and proteins. By engineering human DNA into a host organism. In medicine. Researchers are using it to diagnose and predict disease. and to develop therapies and drugs to treat devastating diseases like cancer. it can be used to make livestock leaner or larger. It is the process of removing DNA from one organism and inserting it into the DNA of another organism. vaccines. giving it new traits. Recombinant DNA Recombinant DNA is one of the core techniques of genetic engineering. Insulin production is an excellent example of the recombinant DNA process. Alzheimer's.MEDICINE S ome of the most promising and powerful applications of genetic engineering are in the field of medicine. that organism can be turned into a factory for important medical products. and cystic fibrosis.fight viral infections several interleukins granulocyte-macrophage colony-stimulating factor (GM-CSF) for stimulating the bone marrow after a bone marrow transplant tissue plasminogen activator (TPA) for dissolving blood clots adenosine deaminase (ADA) for treating some forms of severe combined immunodeficiency (SCID) angiostatin and endostatin for trials as anti-cancer drugs parathyroid hormone 17 . coli. Host organisms can range from bacteria like E. to plants. to animals. Genetically Engineered Pharmaceuticals insulin for diabetics factor VIII for males suffering from hemophilia A factor IX for hemophilia B human growth hormone (GH) erythropoietin (EPO) for treating anemia three types of interferons . Explore more about ways genetic engineering techniques can be used for medical purposes. Recombinant DNA can be used to make crops resistant to pests or disease.
Germline alterations for humans should not be prohibited outright. we recommend a better coordinated effort among relevant regulatory agencies. to ensure there are no gaps in the regulatory framework. 18 . Rejection of this new technology on the ground that it is unnatural or inherently immoral is unwarranted and seems to be based on little more than an instinctive adverse reaction. but with the distinct advantage of producing more predictable and more rapid results. not just with respect to immediate benefits and harms. such as directed breeding. With respect to germline enhancements for plants and animals. such as the Food and Drug Administration and the Department of Agriculture. each proposed alteration needs to be carefully evaluated. its potential benefits outweigh its harms. but also with respect to the effects that the proposed alteration may have on our social structure and the distribution of social goods. given the special risks posed by human germline alterations. However. Enhanced organisms should be rigorously evaluated and tested in isolated conditions prior to their release in the wild. In particular. Biotechnology is an extension of already accepted and well-established techniques.CONCLUSION Genetic engineering has the potential to transform our lives in many positive ways. Legislators and other responsible decision-makers should not implement regulations that unduly restrict implementation of genetic engineering. certainly not in advance of their availability. but provided that it is appropriately regulated. There are risks involved with this new technology. existing mechanisms that ensure the safety of testing protocols should be sufficient for somatic genetic therapies for humans.
Available at http://www.net/content/news/eng/gm-cow-milk-couldrovidetreatment. http://www. Ethical Dangers of Genetic Engineering.php?id=2088.ellendale. http://geneticengineeringmedicine. Epstein.us/hsmain/thoffman/biology/geneticengineeringwebquestpr esentation. Paulo.com/botanical_biotechnology_biology_chemistry/biotechnolog y/genes_genetic_engineering/genetic_engineering_for_human_welfare/biotech_methods _of_dna_profiling.org/wiki/DNA_profiling 11. GM Cow Milk Could Provide Treatment for Blood Disease.nd. 3. Ruiz-Marrero.pdf 6.animalresearch.html 5. http://itotd.com/articles/572/dna-fingerprinting 12. Available at http://www.iptv. 2002. http://www.org/wiki/Genetic_engineering 2.htm 7.eplantscience. http://www.wikipedia.cfm. Available at http://www.php 10.k12.REFERENCE 1.org/wiki/Dolly_(sheep) 19 .wikipedia. 4. http://en. Carmelo.info/en/medical/timeline/Dolly 13. file:///C:/Users/syanmugam/Desktop/Genetic%20Engineering/Applications%20of%20Ge netic%20Engineering. 1999.org/s-r/20/20-01. http://en. Rebelo. http://en.cfm 9. http://www2.corpwatch.scidev.wikipedia. Institute for World Religions.for-blood-disease. Ron.org/article. 2004.com 8.org/exploremore/ge/uses/use2_medical.greens. Genetic Pollution: Biotech Corn Invades Mexico.
http://en.htm 18. http://www.uk/resource/briefings/gm_crops_food.org/english/content/ff_cont3. Pre-U Text STPM Longman Biology Volume 1 and 2 20 .foe.org/wiki/Genetically_modified_food 16.14.com/scitable/topicpage/genetically-modified-organisms-gmostransgenic-crops-and-732 15.co.bionetonline. http://www.wikipedia.nature.pdf 17. http://www.
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