X-gal is a chromogentic substrate of ß-galactosidase. X-gal is used in conjunction with IPTG for detection of ß-galactosidase activity in bacterial colonies in a colorimetric assay to detect recombinants (white) from non-recombinants (blue). X-gal is cleaved at the ß1-4 bond between galactose and the 5-Bromo-4-chloro-3-indolyl part of X-gal by ß-galactosidase via hydrolysis. The cleavage of X-gal results in the production of a water insoluble blue dichloro-dibromo-indigo precipitate at the site of enzymatic cleavage. In cloning strategies with vectors like Lambda-11, M13mp18 and 19, pUC18 and 19, pUR222 the E.coli lacZ gene is transformed to lac cells. After transformation of cells show ß-galactosidase activity in the presence of IPTG and X-gal containing media. The insertion of a DNA fragment into the cloning sites of the lacZ gene results in the disruption of ß-galactosidase activity leading to the appearance of white colonies on X-gal and IPTG containing media. Non recombinant cells produce a blue indifo dye on these media.
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