European Journal of Biotechnology and Bioscience

Authors
Vaibhavi J. Madhad, K. P. Sentheil

Abstract
DNA is the genetic material of majority of all well-defined organisms. In the field of molecular biology, one of the most important procedure is DNA isolation. Genotyping is the process of determining the genes (genotype) of an individual by examining the individual's DNA sequence on the basis of biological assays. There are several different DNA isolation procedures and for choosing the right one depends on the species, the DNA is being isolated from. There is no one “best” procedure because each species’ cells have unique properties that require different methods to release the DNA. In this study, there are different DNA extraction methods to be assessed and to be determined which one is best suited to recover DNA from blood. These procedures does not contain any contaminating agents like proteins & hemoglobins, organic solvents or chaotropic salts like phenol, chloroform, isoamylalcohol & enzymes like Proteinase-K, Pronase that could interfere in the genetic analysis methods to follow like DNA Quantification, RE digestion, Southern Blotting, PCR Amplications, RFLP Analysis & DNA sequencing. The “Alternate Protocols” developed, when compared with the reference protocols employs inexpensive, non-hazardous reagents and yields intact DNA of (30-50µg) from as little as 300 µL of human blood. The Alternate protocol-12 –‘Rapid & Non-Enzymatic method of DNA isolation’- based on the principle of Salting out, completely eliminates the use of hazardous chemicals like Phenol, Chloroform & Isoamylalcohol. It avoids prolonged digestion of samples with Proteinase-K & can be completed within 65 minutes of incubation period. It showed concentration of 100-150 µg of DNA isolated from 1ml of blood & sufficient high purity of 1.5-1.8, suitable for genotyping.