Blue White Selection

Blue white selectionis a widely utilize manner in screening recombinants in cloning. This is based on the factor overlap of lac z gene. The plasmid vectors contain this gene which produces? galactosidaseenzyme. When a gene is inserted mingy tolac z gene, the reading frame allow be distorted and the gene is inactivated. So the change cells will not produce this enzyme and are called skilled cells. After the recombination, the bacterial cells are grown in a mass medium containing X gal (5-bromo-4-chloro-indolyl-? D-galactopyranoside) andIPTG (Isopropyl ? -D-1-thiogalactopyranoside). IPTG acts as the inducer for lac z gene and enhance the production of ? galactosidase. When it is produced, combines with X gal to form a blue glossiness complex called5,5-dibromo-4,4-dichloro-indigowhich is insoluble. The transformed colonies will appear white in colouration and non- transformed cells will appear blue in colour. This method is withal called as insertional inactivation of lac z gene. Hybridizationtechniques are widely used to identify recombinants.This is based on the ability of nucleic acids hybridize with complementary DNA. The transformed cells are transferred on to anitrocellulose membranewhich is subjected to cell lysis. The echo stranded DNA is converted to single stranded DNA and immobilized on the membrane. Then it is treated with radiolabelledprobescomplementary to target DNA. If the desired DNA is present, the probes will be hybridized which can be detected by autoradiography. Apart from these methods,immunochemical methodsare used to detect protein products to screen recombinants.