01:694:215 Lecture Notes - Lecture 10: Repressor, Wild Type, Cloning

• School: Rutgers University
• Department: Molecular Biology and Biochemistry
• Course: 01:694:215
• Professor: A Vershon

Document Summary

Lacz is used as a reporter gene. Blue- cleave x-gal lot of lacz. White- pick up insert does not cleave x-gal no lacz. Yellow- cleave onpg lot of lacz. Clear- pick up insert does not cleave onpg no lacz. Fuse lacz with a gene that we are interested in. Mutational analysis of repressor and activator proteins and their binding sites. Genetic analysis- make mutations and see what happens. Mutant protein- mutate the activator or repressor protein. Mutant site- mutate the site where the activator or repressor attaches. If mutant increases expression the element is a repressor. If mutant decrease expression the element is an activator. Site directed mutagenesis- make changes extend primer synthesize new strand transform bacteria. Recognizes that the base pairing is wrong. Either fixes the wild type or mutant. Endogenous- mutating the sites in the context of their natural promoter. Introduce mutations in the promoter region and test expression.