A Textbook Of Biotechnology

Robert Hook (1655) was the first person who observed cells of cork (dead cells) under the microscope, which appeared like honey comb. Leeuwenhock (1674) observed white blood cells with the help of a microscope with some organization inside. Schleiden and Schwann (1838–39) proposed the cell theory, according to which the cell is the unit of structure of living system (cell is the unit of life), i.e., all living organisms are made up of cells (except for viruses, which are living but have acellular organization and are composed of only proteins and DNA/RNA, some viruses also contain membranes) and the cell arises from the pre-existing cells.

For various living organisms, 16 macronutrients and 11 micronutrients (total 27) are essential. In living organisms, H, O, C of N are most abundant elements, constituting about 99% of mass, where C = 50–60%, O = 25–30 and N = 8–10%.

Biomolecules are mostly organic compounds, e.g. nucleotides, nucleic acids, amino acids, proteins, lipids, carbohydrates, vitamins, hormones, etc.; a few are inorganic, e.g. H2O, salts, mineral ions, etc. Some are small molecules (micromolecules), e.g. minerals, water, amino acids, sugars (mono and disaccharides), lipids, nucleotides, whereas, others are larger in size (macromolecules), e.g., proteins, nucleic acids, polysaccharides.

The word lipid has been derived from the Greek word ‘lipos’ and they are the esters of fatty acids with alcohols. Generally, they are triglycerides, containing three fatty acids linked to the alcohol, glycerol. Lipids are hydrophobic and may contain some hydrophilic groups. They are made up of C, H and O, but sometimes P, N, S are also present. They are insoluble in water (polar solvent) but, soluble in organic solvents (non-polar solvents), e.g. benzene, petroleum ether, acetone, chloroform, hot alcohol, etc.

For growth and reproduction, different types of cells undergo division process by various ways. Thus, the prokaryotic cells simply divide through binary fission method, in which the parent cell divides into two identical daughter cells for both growth as well as asexual reproduction. Here, after DNA replication, the two DNA molecules get distributed preciously into two daughter cells with the help of mesosome (an invagination of cell membrane). Unlike prokaryotes, the eukaryotic cells divide by two types of more complex methods of cell division:

The term ‘genetics’ was coined by W. Bateson (1905) and is the science that deals with heredity and variation. Heredity is the transmission of traits from parents to offspring, whereas variation can be hereditary (that are transmitted from generation to generation and arising mainly due to independent assortment of chromosomes and recombination in sexual reproduction and to some extent due to mutations) or environmental (that do not get transmitted to next generation).

Nucleic acids are long chain polymers (actually macromolecules as they consist of four types of nucleotides and they are not repeated in an orderly manner) of nucleotides (monomers). Each nucleotide is made up of a sugar (either ribose or deoxy-ribose), a base (thymine, cytosine, uracil, adenine or guanine) and a phosphate group (where the sugar and the base together constitute a nucleoside).

Genes (DNA) are generally expressed through transcription (transfer of information from DNA to RNA, e.g., synthesis of mRNA, tRNA, rRNA) and translation (transfer of information from mRNA to proteins) of language of nucleic acids (mRNA) into language of proteins. Thus, the sequence of bases in DNA and RNA determine the sequence of amino acids in proteins. Some genes do not contain any information of proteins and are expressed only through transcription, e.g., tRNA, rRNA genes. This one way flow of information from DNA to RNA and finally to proteins is known as central dogma

Acorrelation exists between sequences of nucleotides in DNA (or mRNA) and amino acids in the proteins synthesized, and this relationship is called ‘Genetic Code’ .

Mutations are the abrupt changes in genetic material that alter the chemical structure of genes at molecular level (gene mutations or point mutations). Mutations occur more frequently in certain regions of DNA that are called hot spots of mutations. The first mutant was isolated by Morgan (1910), that was a white-eyed male Drosophila melanogaster (fruit fly, where the normal eye colour is red) and this character was located on the X-chromosome. Thereafter, about 500 mutants were discovered in Drosophila.

In prokaryotic cells, only one type of RNA polymerase synthesizes all the three types of RNAs (mRNA, tRNA and mRNA) and, since, the DNA is not bounded by any nuclear membrane, transcription (formation of mRNA) and translation (formation of proteins) processes occur simultaneously in the same cytoplasm. (In eukaryotes, transcription or formation of mRNA occurs in nucleus, whereas, translation or protein synthesis in cytoplasm).

The technology of production of recombinant DNA by inserting an isolated or artificially synthesized fragment of DNA at a desired place into a vector and then introduction of this recombinant DNA into a host cell is known as genetic engineering (recombinant DNA technology), e.g., transfer of human insulin gene into Escherichia coli where it expressed to produce the insulin protein that is used for the treatment of diabetes. Thus, a single copy of a gene can be cloned into an indefinite number of copies, all being identical, by multiplication of the bacterium or phage containing that particular gene in the usual manner. This technique is also known as gene cloning. Genetic engineering in eukaryotes requires new methods and tools as, unlike in prokaryotes, the chromosomes are found in the nucleus and, moreover, many genes are split genes (exons interrupted by introns) that cannot be correctly expressed when cloned in prokaryotes (prokaryotes do not have machinery for the processing of HnRNA, and when cloned as such in prokaryotes, the entire HnRNA is translated into respective protein).

Biotechnology includes all industrial processes mediated by living organisms at some step or the other. Microorganisms can be exploited for the production of industrially important biochemicals, like, various enzymes, pharmaceuticals, hormones, antibodies, which were earlier available only by sacrificing the animals. Biotechnology is also being used for a variety of crop plants and domestic animals improvement programmes throughout the world.

All biotechnological processes are performed within bioreactors (may be a culture vessel, an open tank or sophisticated fermenters) containing correct medium provided with optimum growth conditions, like pH, temperature, aeration, light (for photosynthetic organisms), etc., The growth of the organisms is the increase of cell material that can be measured in terms of many parameters, like mass (dry weight or fresh weight), total amount of proteins, photosynthetitic pigments, number of cells, etc., Doubling time is the period required for doubling the biomass (due to cell division as well as due to cell growth) which varies from organism to organism, e.g., for bacteria it is about 0.25–1.00 h, for yeast 1–2 h, for plant cells 20–70 h and for animal cells 15–48 h (generation time is the time period required for doubling of the cell number due to cell division).

Food industry is the largest industry in the world and food biotechnology includes improvement in taste, consistency, colour, nutrition, safety and preservation of the food. Developments in food preservation methods have made many of the seasonal foods to be available all the year round. Fruits, vegetables, cereals, meats, etc. require some degree of processing, and the relatively bulky raw agricultural food products are transformed into stable, convenient and palatable foods and beverages.

The term enzyme was given by F.W. Kuhne (1878). They are generally made up of proteins and catalyze biochemical reactions. They are specific and have three-dimensional structure with highly specific one or more active sites for the recognition of substrates. The active site (lock) of the enzyme binds with its substrate (key) as "lock and key model", given by Fisher

Continuously increasing population is leading to the intensive agriculture (to get greater amount of food and other products on the same limited land by the use of improved varieties of crop plants, which require higher amount of chemical fertilizers, insecticides, herbicides, etc.), urbanization, industrialization, deforestation, and excessive utilization of various resources (fossil fuels, underground water, etc,). This leads to the degradation of our environment by generating various kinds of liquid, solid, gaseous and hazardous wastes, particularly in the developed countries, where, only about 15% population consumes about 85% resources to generate about 85% of the wastes (most of which are non-biodegradable) in comparison to the developing countries. On the other hand, in developing countries higher population, limited technological knowledge and finance facilities for the treatment of wastes are the major causes of environmental degradation. Earlier, when the population was small the environment was sustainable and the biodegradable wastes generated were easily decomposed by the decomposers. But now, the amount of waste generated is so great that the limited natural capabilities of the environment are unable to decompose it properly, particularly the toxic non-biodegradable ones, leading to the accumulation of such wastes in food chains (biomagnification of insecticides, like, dichloro diphenyl trichloro ethane or DDT, herbicides, etc.) as well as in the surroundings. Large-scale production and application of synthetic chemicals in most industrialized countries is a problem of serious concern causing serious health hazard. Organic chemicals that cannot be easily degraded by microorganisms or totally resistant to biodegradation are known as recalcitrants, e.g., lignin; whereas, synthetic compounds (foreign substances) not formed by natural biosynthetic processes are called xenobiotic compounds (that can be recalcitrants) and often have toxic effects, e.g., insecticides DDT, BHC (benzene hexa-chloride), etc.

The world depends for its energy requirement upon three major fossil fuels—coal, natural gas and oil, which would become exhausted completely in near future (coal may last for 200 years, natural gas and oil for 40 years). Realizing energy crisis during 1970s when the oil prices increased massively and due to environmental awareness, the scientists are trying to find the alternative non-conventional cleaner sources of energy, like wind, solar, tidal and wave, geothermal and bio-energy. Photosynthetic organisms convert dilute solar energy into more concentrated chemical energy (e.g., carbohydrates) with 3–4% efficiency, making the biomass (2 × 1011 tonnes C fixation/ year). Photosynthetically derived biomass (which does not have much calorific value), like forest, agricultural and animal residues and domestic organic wastes can be converted by fermentation into clean renewable fuels (like alcohol and CH4) possessing quite higher energy value. With time, continuous depletion of fossil fuels is making them increasingly expensive and in such situation biofuels may become more economic. Biofuels can be used for cooking, generation of electricity or in automobile engines. The CO2 released during their combustion is entrapped in the photosynthesis (which also produces O2) for biomass production and, thus, do not damage the environment as the fossil fuels do. Except for biogas, other biofuels are not economical.

Human is totally dependent on agriculture and many countries are still not self-sufficient in food production. Biotechnology is creating a revolution in this field to provide higher quality and lower cost products to meet the requirement of continuously increasing human population. Since early times, man is trying to improve the quality and productivity of agriculturally important plants by selection of better varieties and employing traditional breeding programmes involving sexual crosses, which are slow and difficult processes. Extensive research is also going on with forest trees, as forest resources are continually depleting fast because of indiscriminate deforestation. Trees have long generation times and desired traits can be achieved by faster tissue culture and genetic engineering technologies.

About 200 years ago the life was very harsh and less than 2% people were able to live beyond the age of 65 years. With research, it became possible to control various diseases by using different antiseptics, vaccines, antibiotics, etc. Life expectancy over the last 150 years rapidly increased from 35 to almost 80 years. Today, the infectious diseases are no longer the main threat to life and only the chronic diseases, like cancer, cardiovascular diseases, etc. are dangerous and can be solved through genetic manipulation that has lead to the development of new strains with higher productivity and new medicines. New techniques of protoplast fusion and gene transfer technologies gave products, which resulted in the decrease in cost of production.