Practice question

Successful blue-white selection requires three supplements to Luria-Bertani agar besides routine nutrients for bacterial growth. Ampicillin ensures plasmid retention by selecting bla-positive transformants and inhibiting cell wall synthesis in plasmid-free segregants. Isopropyl β-D-1-thiogalactopyranoside, a non-hydrolyzable lactose analog, derepresses lac operon by inactivating LacI repressor, inducing strong expression of plasmid-borne lacZα. X-gal provides chromogenic substrate; cleavage by β-galactosidase yields insoluble blue precipitate. Combined medium permits simultaneous positive selection for plasmid presence and colorimetric screening for insertional inactivation, distinguishing recombinant white colonies from non-recombinant blue colonies.