What is Genetic Engineering?
Genetic engineering, recombinant DNA technology, genetic modification or manipulation (GM) and gene splicing are terms that apply to the direct manipulation of an organism’s genes.
The term “genetic engineering” was coined in Jack Williamson’s science fiction novel Dragon’s Island, published in 1951, two years before James Watson and Francis Crick showed that DNA could be the medium of transmission of genetic information.
Later on Arber in 1950 discovered enzymes that degrade bacterial viruses and Smith in (1970) purified the enzymes and characterized them
The founding fathers of genetic engineering were Stanley Cohen and Herbert Boyer who in 1973 in vented the technique of DNA cloning
STEPS INVOLVED IN SEQUENCING
Separation of DNA
In this step, preparation of genomic DNA from the cell line of the desired organism is done.
Fragmentation
Fragments of DNA are obtained using various types of restriction enzymes. This is due to the presence of various restriction sites in the sequence.
Polymerization
The fragments to be sequenced are cloned in a vector plasmid or cosmid (to get sufficient quantity of fragments to be sequenced). Polymerization of the DNA sequence can be done by using a PCR naturally or artificially.
Labeling
The fragments are then labeled according to the requirement. It might be radioactive labeling or fluorescent labeling. The labeling is done basically for identification purposes.
Sequencing reactions
Many of the methodologies of sequencing have been used. For e.g Maxam Gilbert sequencing, Sanger’s chain termination methods etc . The later method is widely used and is the base of modern sequencers.
Gel electrophoresis
The colonies of DNA fragments are run in the gel to identify and infer the results. Four different lanes for different nucleotides are made. Analysis in modern way are done in capillary electrophoresis unit.
Visualization
The fragments are now visualized using staining method, and fluorescent readings.
Computer analysis
The overlapping regions of the fragments are determined using computer analysis. The overlapping sequences are assumed into continuous sequence. This is a base method in modern sequencers which are completely computerized.
Annotation
Conquering the pieces of DNA with known sequences and database which helps in annotation and identification of unknown gene.
Further reading:
An animation of sanger’s method for DNA sequencing
How do we obtain the graph?
For my notes and detailed procedure of DNA click HERE
Like this:
Filed under: genetic engineering
