Question

Give an account of the Blue-White Method of selection of recombinants

Answer 1

Recombinant DNA (rDNA) technology is the technique of manipulating the genome of a cell or
the organism so as to change the phenotype desirably

Following are the basic steps involved in the process:

1) Isolating genomic DNA of a ‘donor’. The cell or organism from which the required gene is taken is called ‘donor’

2) Fragmenting this DNA using “molecular scissors’ (Enzymes): Different enzymes used are; restriction endonucleases, DNA ligase, reverse transcriptase, DNA polymerase, alkaline phosphatases etc. The restriction endonucleases are used to cut DNA at specific points. They are called biological / molecular / chemical scissors / knives / scalpels.

3) Screening the fragments for a ‘desired gene’

4) Inserting the fragments with the desired gene into a ‘cloning vector’ - Vectors are the DNA molecules used to transfer genetic material into another cell. (a plasmid, cosmid or phage DNA), so as to develop a recombinant DNA or chimeric DNA.

5) Introducing the recombinant vector into a competent host cell.

6) Culturing these cells to obtain multiple copies or clones of the desired fragment of DNA.

7) Using these copies to “Transform” suitable host cells so as to express the desired gene.

8) Selection of transformants by the blue-white method of selection of recombinants.

It is a screening technique that allows for rapid and convenient detection of recombinant bacteria in vector – based molecular cloning experiments. Recombinant DNA is inserted into a competent host cell viable for transformation, which is then grown in presence of X – gal. The cell transformed with vectors containing Recombinant DNA will produce white colonies, cells transformed with non – recombinant plasmids (i.e. only the vector) grow into blue colonies. This method of screening is usually performed using a suitable bacterial strain, but other organisms such as yeast may also be used.