After the recombinant is introduced into the recipient cells, it can be used for further screening and identification after a short period of culture expansion. Screening refers to the process of identifying positive grams (recombinants) of recombinant DNA molecules from transformed cell populations or gene libraries through specific methods. The selected positive clones can obtain a large number of copies of the required gene fragments through multiplication, and through further identification, their correctness can be finally determined, laying the foundation for their subsequent research.
In addition to antibiotic resistance screening, blue-white screening is also a common method in molecular cloning: when foreign DNA is inserted into the vector LACZ-tagged gene, resulting in inactivation of the -galactosidase gene, the color change of E. coli transformant colonies in medium supplemented with X-gal-IPTG can be directly observed.
Galactosidase hydrolyzes lactose into galactose and glucose, and when host cells are cultured in a medium supplemented with X-Gal (5-bromo-4-chloro-3-indole-D-galactoside) and the lactose inducer IPTG (isopropyl--D thiogalactoside), the functional galactosidase formed by gene complementarity can cleave the colorless X-gal in the medium into galactose and the dark blue substrate 5-bromo-4-chloro-indigo, causing the colonies to exhibit a blue reaction. However, when the exogenous DNA is inserted into the multiple cloning sites inside the vector lacz-tagged gene, the code-reading structure of the -galactosidase gene can be blocked, and the peptide encoded by it becomes inactive, resulting in a white colony. Thus, based on this chromogenic reaction of galactosidase, recombinant clones containing foreign DNA insertions can be detected.