MCB 2050 Lecture Notes - Lecture 15: International Hapmap Project, Avidin, Embryonic Stem Cell

• School: University of Guelph
• Department: Molecular and Cellular Biology
• Course: MCB 2050
• Professor: D Mosser

Document Summary

Mcb 2050 - basic concepts of each unit. Unit 1. 1: basic techniques used to clone genes. Cloning steps: digest vector and dna of interest, anneal-ligate, transform: coli, selection. Restriction enzymes: cut specific sites in dna by breaking phosphodiester bonds leaving either blunt or cohesive ends. function in bacteria: degrade invading dna, Bacteria must methylate their own dna to prevent digestion by their own restriction enzymes (re). complimentary cohesive (or blunt) dna ends created by re digestion can be ligated together with dna ligase. 3 essential components: origin of replication, dominant selectable marker (antibiotic resistance gene), unique re sites for gene insertion screening for recombinant plasmids in the pbluescript cloning vector: Spread transformants on agar with x-gal: blue colonies (lacz intact no insert), white colonies (lacz gene disrupted insert present). eukaryotic expression vectors: Bacterial ori to propagate the plasmid and selectable marker (ampicillin) for selection of bacteria that contain the plasmid after transformation eukaryotic promoter,