BIOL309 Study Guide - Midterm Guide: Puc19, X-Ray Crystallography, Nylon

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28 Dec 2011
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Module 6: cloning vectors vector: nucleic acid molecule used to transport and replicate sequences of interest into a host organism. Lacz expression is induced by growth on iptg (isopropyl-beta-d- thiogalactopyranoside) and turns blue in the presence of xgal. Cacl2 affects the cell wall, allowing dna binding, heat shock stimulates dna uptake. Wash with ice cold water (cold prevents cell growth) to remove electrolytes. To test competency, a known amount of supercoiled plasmid is added ex. add 5ng of puc18 to 0. 1ml of cacl2-treated e. coli cells with 1ml lb broth and plate 50ul and 5ul of transformants on selective media. 137 colonies found on 50ul and 18 on 5ul plate. 0. 1ml cells + 1ml broth = 1. 1ml total. 22 plates x 137 colonies = 3x10^3 transformants. If 5 ng of dna produces that many colonies, how much would 1ug dna produce? (3x10^3)/0. 005ug = (x/1ug) x=6x10^5 transformants improving transformation efficiency modify salt solution (add salts like mg to.

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