Genetic engineering principles and applications

Genetic Engineering is a tool in the hands of man to break the species barriers to create a more productive and controllable world. This is a delicately balanced issue and unless we exercise enough restraint and responsibility we may end up endangering ourselves and all other forms of life.

Genetic Engineering is the science of gene manipulation. Genetic information is specific to each and every organism in the world. Genetic Engineering is in effect the science that deals with the controlling of the expression of the individual genes within a cell. Today the advancements in science have made possible the selective study of the individual segments of the DNA of a particular species, to isolate them and to infuse them in the DNA of a totally different organism. Genetic Engineering can be viewed as a breakthrough in the study of organisms that effectively disintegrates the distinctions that exists in the different species of the world. The advancements in recombinant DNA technology have expedited the research in genetics. Let us analyze this subject in a little detail and in the process analyze its pros and cons and its future implications.

Importance of Genetic Engineering

Since 1980 when the first artificially synthesized Insulin was made available for treating patients the technique of coercing the microbes to synthesize a variety of useful substances of medical value has attracted the attention of microbiologists all over the world. Today genetically engineered microbes continue to produce a wide variety of substances like interferon, vaccines, growth hormones, etc. With this remarkable breakthrough in cell biology scientists are for the first time vested with the keys to unravel the mysteries of complex process of life. Genetic engineering is a tool in the hands of the researchers to manipulate the genes and induce the cells to produce substances that they never made before. Apart from the medicinal value there are some ethical issues to be considered and genetic engineering has become a debatable topic over the last few years with scientists vying with each other to alter the basic code of life.

The first and the important step in creating a genetically engineered bacterium or any other host is to isolate the gene from the cell. All human cells share the common genetic information between them but the difference lies in the fact that the expression of the genes that are responsible for the synthesis of the specific proteins is different and is related to the environment of the cell. John Sulston, former director of the Sanger Centre says, "You cannot define the function of genes without defining the influence of the environment. The notion that one gene equals one disease, or that one gene produces one key protein, is flying out of the window." [Venter Craig]. It is this selective expression property of the gene that is used by scientists to identify and trace down the individual genes.

Genes Expression and Cell Behavior

Gene manipulation is defined variedly in different countries. "The formation of new combinations of heritable material by the insertion of nucleic acid molecules, produced by whatever means outside the cell, into any virus, bacterial plasmid or other vector system so as to allow their incorporation into a host organism in which they do not naturally occur but in which they are capable of continued propagation" is how it is defined in the United Kingdom. [OLD.R.W, 1]. The genes in fact directly control and regulate all the natural body processes. Different cells in the body perform different actions and this is due to the selective expression of the genes. For example cells in our brain do not produce insulin while those in our liver produce it. Similarly bone marrow cells produce Red Blood Corpuscles while liver cells don't do the same. To understand this peculiar cell behavior we have to look into the process that takes place in the cell that either activates the gene or deactivates it.

The key to understanding the process of gene expression is to study the mRNA molecules. This is so because cells that promote the expression of a gene have the mRNA molecules within them. The gene constructs a mRNA molecule whenever the particular protein that it creates is needed by the cell. Extricating the mRNA molecules from the other constituents of the cell is an intricate process in itself but it is a lot easier than identifying the different strands in the chromosome. Usually a centrifuge performed on the cell constituents separates the different molecules. These isolated mRNA molecules contain the genetic information required for the synthesis of the corresponding protein molecule. Now that the mRNA is isolated the next step is to transfer this back in to the specific DNA sequence.

Over the years this particular area has been the bottleneck for scientists involved in genetic engineering. However the answer was found and the use of enzymes effected the trick. This method was first detected in viruses. The essential idea is to use enzymes that convert mRNA into DNA. These enzymes are called as reverse transcriptases because they reverse the process of transcribing DNA into mRNA. Thus adding reverse transcriptase enzymes to human mRNA strands will create Human DNA strands. Finally some other enzymes like the DNA polymerase used to convert the single strand DNA to its original double helical structure thereby completing the process of synthesizing the complementary DNA (cDNA an exact replica of the mRNAs). [Chhatwal G.R, 1998,99]

Protein (the Building Block)

Proteins are fundamental building blocks of the cell. They are nothing but different combinations of Amino acids and different proteins perform totally different functions within the human body. Hormones, Enzymes and antibodies are all different kinds of protein molecules. The synthesis of proteins in the cells is achieved by means of the genes. Each gene is actually a specific segment of the DNA having instructions to create a particular type of protein upon request. Request for the synthesis of proteins is again controlled by what are known as promoters that are unique for each gene. It is the function of these promoters to either activate or deactivate the expression of the individual genes and hence the production or not of the corresponding proteins. All genetic engineering techniques are invariably concerned with manipulating and controlling these promoters and hence the gene behavior. [Synthesis/Regeneration Magazine]

Gene Cloning

Gene cloning constitutes the core of genetic engineering. By means of cloning scientists are able to develop an exact replica of the gene or the all-important proteins that they produce naturally. Usually the following steps are involved in a cloning process. Firstly the reguired gene has to be extricated from the chromosomes and for this scientists use certain enzymes that would facilitate the breakage of the bonds between the different strands. In the genetic parlance these enzymes are called as the 'restriction enaonucleases'. There is also another way to create the gene artificially by using what is called as the gene machine. The next step in the process is to insert the extricated gene into a vector so that the gene can be accepted by the host such as a bacteria or virus. Once this is done these microbes can be cultured to get multiple copies of the genes. [Chhatwal G.R, 1998, 88]

The procedure of cloning however is much more complicated and involves refinement at every stage of the process. Genetic engineers refer to this as 'enrichment technique'. Invaluable medicinal substances (Insulin) can be synthesized by genetic engineering. Vectors can be used to incorporate special feature in the clones and to restrict the growth of all other hosts which do not have this property. For example we can choose to incorporate the property of drug resistance in the host and in order to selectively isolate them or purify them we can culture the bacteria in a medium of antibiotics. This way only those hosts that are incorporated with the special genes will grow and so we obtain an unadulterated and pure bacterium with drug resistance properties.

Plasmids as Vectors

Plasmids are nothing but small circles of DNA inside the bacterial cell. They are present outside the chromosome and hence are called extra chromosomal. Generally Plasmids are double stranded circular DNA molecules. Today Plasmids are considered as the ideal cloning vehicles. The important properties to look for in a Plasmid (as a cloning vehicle) are that they must have low molecular weight and have the ability to infuse the phenotypic traits on the host cell. [OLD.R.W, 46]. Another natural and inherent property of the Plasmid is that it can readily pass on from one cell to the other. This property of the Plasmid to readily enter into any cell is very important for Genetic engineering. This allows scientists to extract Plasmids from the bacterial cell and 'Stitch' the cDNA into it. Once this is done the Plasmid by its natural ability to penetrate into the cell finds its way back into the bacteria.

Enzymes are again the key to whole process called 'Stitching'. Restriction enzymes are very meticulous in their purpose and they do their function very precisely. Today there are more than 300 restriction enzymes and the list ever continues to grow. Each of these restriction enzymes precisely identify and target particular DNA bases. When treated with DNA the restriction enzymes scan through the DNA strands and identify the specific sequence bases and cut through them. This opening makes possible the insertion of the human gene. This Plasmid-human gene combination is what is called as the recombinant Molecule. There is one more step to be done after the recombinant DNA is introduced into the bacteria. This is to make the bacteria express the newly introduced gene. This is achieved by tagging the bacterial control regions to the human gene. Then the bacteria pickup the signal and create the mRNA. [Chhatwal G.R, 1998, 101]

Extracting the Proteins

Even though we have managed to create the vital human proteins inside the bacterial cell we still need to extract them in a pure form from the other cell constituents. This is called as the 'Down Stream Processing' in the genetic engineering and biotechnology parlance. There are many techniques employed in the isolation and separation of the protein form the cell. Genetic engineers are continuously refining their techniques to ease this 'Down-stream processing'. So far we have discussed the actual technique of genetic engineering or gene manipulation. Let us now look into the application areas.

Applications of Genetic Engineering

Medical Applications

There are hundreds of genetic disorders that are caused by mutations. For most of these diseases there is no known cure. Gene therapy is already a reality and may soon prove to be the panacea for all genetically induced diseases. The advancements in the area of Biochips technology have greatly altered our ability to treat diseases and genetic disorders in particular. Andrei Mirzabekov, one of the leading biologists researching at the Argonne and Engelhardt says, "Instead of reading DNA one letter or word at a time, they read whole phrases and sentences at a time," "By combining biochips with robots and computers," Mirzabekov further adds on that, "we can find one genetic variation among three billion DNA bases in a matter of minutes. Conventional methods take days." [TheHostPros]

Gene chips are nothing but silicon wafers in which the DNA strands are itched out using high precision nano technology. By using these gene chips we can search and identify unknown strands in quick time. Currently genetic disorders, genetically modified agricultural products and bio medical researchers are the main users of gene chips. [Arthur Chiou] Many leading companies like Motorola and Packard are already venturing into mass production of biochips and the technology required for analyzing the biochips. Richard McKernan, president of Packard Instrument Company says, "With a commercial biochip to rapidly and economically perform genetic analysis, within a few years we should see better pharmaceuticals developed more rapidly, faster and more accurate medical diagnostics, a heightened ability to assess and possibly repair environmental damage, and better, more hardy, and healthier crops," [TheHostPros] So we can be sure and hopeful that in the near future medical diagnosis can be expedited and in many cases genetically caused diseases can be prevented.

Agricultural Sector

Genetically modified food is the hot topic of the day. Vitamin A deficiency has been the main cause for blindness and even death in thousands of people especially in the under developed nations. Some scientists argue that genetically modified rice (with added Vitamin A) will be the solution to this problem. While all this is good to hear there are many experts who opine that GM (Genetically Modified) Crops are a serious problem. "Because gene containment is next to impossible with the current generation of GM crops, this discriminatory stance (i.e., regulations based on "process rather than products") has led to several international 'incidents' over the past few years." [Nature Biotechnology, June 2002,527] While genetically modified food materials are gaining rapid acceptance within the United States many European nations are still skeptical about the GM products.