pGLO Tutorial - English

pGLO™ Transformation and Purification of Green Fluorescent Protein (GFP)

[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],Instructors

Why Teach Bacterial Transformation and Protein Purification? ,[object Object],[object Object],[object Object],[object Object],[object Object]

pGLO ™ Bacterial Transformation Kit ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

Green Fluorescent Protein (GFP) Chromatography Kit ,[object Object],[object Object],[object Object],[object Object],[object Object]

Workshop Time Line ,[object Object],[object Object]

Central Framework of Molecular Biology DNA RNA Protein Trait

Links to Real-world ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

Using GFP as a biological tracer http://www.conncoll.edu/ccacad/zimmer/GFP-ww/prasher.html With permission from Marc Zimmer

pGLO Bacterial Transformation Kit

Transformation Procedure Overview ,[object Object],Day 1 10

What is Transformation? ,[object Object],GFP Beta-lactamase Ampicillin Resistance

What is a plasmid? ,[object Object],[object Object],[object Object],[object Object]

Bacterial DNA Plasmid DNA Bacterial cell Genomic DNA

The Many Faces of Plasmids Scanning electron micrograph of supercoiled plasmid Graphic representation

Gene Expression ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

Bacterial Transformation Beta lactamase (ampicillin resistance) pGLO plasmids Bacterial chromosomal DNA Cell wall GFP

Transcriptional Regulation ,[object Object],[object Object],[object Object]

Transcriptional Regulation B A D araC B A D araC RNA Polymerase Effector (Arabinose) araC B A D ara Operon RNA Polymerase Z Y A Z Y A LacI Effector (Lactose) Z Y A LacI lac Operon

Gene Regulation RNA Polymerase araC ara GFP Operon GFP Gene araC GFP Gene araC GFP Gene Effector (Arabinose) B A D araC B A D araC RNA Polymerase Effector (Arabinose) araC B A D ara Operon

Methods of Transformation ,[object Object],[object Object],[object Object],[object Object]

Transformation Procedure ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

Reasons for Performing Each Transformation Step? ,[object Object],[object Object],[object Object],Ca ++ Ca ++ O CH 2 O P O O O Base CH 2 O P O O O Base OH Sugar Sugar O Ca ++

Why Perform Each Transformation Step? 2. Incubate on ice slows fluid cell membrane 3. Heat-shock Increases permeability of membranes 4. Nutrient broth incubation Allows beta-lactamase expression Beta-lactamase (ampicillin resistance) Cell wall GFP

What is Nutrient Broth? ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

Grow? Glow? ,[object Object],[object Object],[object Object],LB/Amp LB/Amp/Ara LB

Webinars ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object]

pGLO Tutorial - English

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pGLO Tutorial - English