Biotechnology is the science of applied biological process in which there is a controlled use of biological agents such as microorganisms or cellular components for beneficial use. A Hungarian Engineer, Karl Ereky (1919) coined the term biotechnology. Biotechnology broadly categorized into traditional practices and modern practices. Traditional biotechnology includes our ancient practices such as fermentation. Single Cell Protein (SCP) organisms are grown in large quantities to produce goods rich in protein, minerals, fats, carbohydrates and vitamins. The modern biotechnology embraces all the genetic manipulations. The recombinant DNA technology is a technique of modern biotechnology in which transfer of DNA coding for a specific gene from one organism is introduced into another organism using specific agents like vectors or using instruments like electroporation, gene gun, liposome mediated, chemical mediated and micro injection. Other tools are enzymes and host organisms. The enzyme restriction endonuclease is a molecular scissor that cleaves DNA into fragments at or near specific recognition sites with the molecule known as restriction sites. Other enzymes are DNA ligase and alkaline phosphatase. DNA ligase enzyme joins the sugar and phosphate molecules of double stranded DNA. Alkaline phosphatase is an enzyme which adds or removes specific phosphate group of double stranded DNA.
A vector is a small DNA molecule capable of self replication and used as a carrier of DNA inserted in the host cell. Few examples of vectors are plasmid – pBR 322, cosmid – Lambda phage, M13, Phagmid , BAC, YAC, transposon, shuttle vector and expression vector.
After production of recombinant DNA molecule has been generated is introduced into a suitable host cell. Type of host cell depends upon the cloning experiment. E.coli is the most widely used host organism. There are two kinds of gene transfer methods in plants. They are direct or vectorless gene transfer and indirect or vector mediated gene transfer. Direct gene transfer includes chemical mediated gene transfer, micro injection, electroporation. Gene gun method and Liposome mediated method of gene transfer. Indirect or vector mediated gene transfer is a method of gene transfer with the help of a plasmid vector. In this method Ti-plasmid from Agrobactirum tumefeciens has been used extensively for vector mediated gene transfer.
After the introduction of rDNA into a host cell, it is essential to identify those cells which have received the rDNA molecule. This process is called screening. One of the method of recombinant screening is blue white selection method Replica plating technique in which the pattern of colonies growing on a culture plate is copied. Electrophoresis is a separating technique used to separate different biomolecules.
Blotting techniques are widely used tools for identification of desired DNA or RNA fragments from larger number of molecules. Some of the genetically modified crops are herbicide tolerant – Basta, Dhara mustard, insects resistance – Bt crops, flavrSavr – Tomato, Golden rice. Biopolymers are polyhydroxybutyrate (PHB), polylactic acid (PLA) and green fluorescent protein (GFP) is used to make biosensors. Other applications are biopharming, bioprospecting, biomedication and biofuel, etc.
3’ Hydroxy end: The hydroxyl group attached to 3’ carbon atom of sugar of the terminal nucleotide of a nucleic acid.
Bacterial artificial chromosomes (BAC): A cloning vector for isolation of genomic DNA constructed on the basis of F-factor.
Chimeric DNA: A recombinant DNA molecule containing unrelated genes.
Cleave: To break phosphodiester bonds of dsDNA, usually with a restriction enzyme.
Cloning site: A location on a cloning vector into which DNA can be inserted.
Cloning: Incorporation of a DNA molecule into a chromosomal site or a cloning vector.
Cloning Vector: A small, self-replicating DNA inserted in a cloning gene.
COS sites: The 12-base, single strand, complementary extension of phage lambda (l) DNA.
DNA Polymerase: An enzyme that catalyses the phosphodiester bond in the formation of DNA.
Endonucleases: An enzyme that catalyses the cleavage of DNA at internal position, cutting DNA at specific sites.
Genome: The entire complement of genetic material of an organism.
Insert DNA: A DNA molecule incorporated into a cloning vector.
Ligase: An enzyme used in genetic engineering experiment to join the cut ends of dsDNA.
M-13: AssDNA bacteriophage used as vector for DNA sequencing.
Phagemid: A cloning vector that contains components derived from both phage DNA and plasmid.
Plasmid: Extrachromosomal, self-replicating, circular dsDNA containing some non-essential genes.
Restriction map: A linear array of sites on DNA cleaved by various restriction enzymes.
Shuttle Vector: A plasmid cloning vector that can replicate in two different organisms due to the presence of two different origin of replication OriEUK and OriE. coli
Taq polymerase: A heat stable DNA polymerase isolated from a thermophilic bacterium Thermus aquaticus.
Vectors: Vehicles for transferring DNA from one cell to another.
Biofuel: Fuels like hydrogen, ethanol and methanol produced from a biological source by the action of microorganisms.
Bioleaching: Process of using microorganisms to recover metals from their ores or contaminant environment
Bioremediation: Process of using organisms to remove or reduce pollutants from the environment.
Green Technology: Pollution-free technology in which pollution is controlled at source.
Phytoremediation: Use of certain plants to remove contaminants or pollutants from the environment (soil, water or air).
Recombinant: Cell / Organism formed by a recombination of genes.
Transformation: Process of transferring a foreign DNA into a cell and changing its genome.
Vector: Agent used in recombinant DNA technique to carry new genes into foreign cells.
Wild Type: Natural form of organisms.