PGLO Transformation LAB AP LAB 7 - Brookings …：pGLO转化实验室AP实验室7 -布鲁金斯….ppt

Transformation Results Cells with pGLO plasmid GROW GLOW-can grow on media with antibiotic GLOW on media with arabinose (turns on GFP gene) LB/AMP/ARA PLATELuria Broth+ antibiotic|+ arabinose + + PGLO = Plasmid added → +pGLOLB/amp +pGLOLB/amp/ara -pGLOLB/amp -pGLO LB /faculty/michael.gregory/files/Bio%20101/Bio%20101%20Laboratory/Bacterial%20Transformation/results.htm * * * * * * * * * * * * * * pGLO Transformation LABAP LAB 6 http://www.mshri.on.ca/nagy/GFP%20mice.jpg BIO-RAD lab book pGLO ori bla GFP araC Aequorea victoria: Source of “glowing gene” for this experiment Jellyfish Gene put into Other Critters /files/21291/monkey_x600.jpg http://www.lafuga.de/GFP_pig.jpg PLASMID Extrachromosomal DNA Often carry genes for antibiotic resistance Can be passed from one bacterium to another /~owens/age2062/OnLineBiology/OLBB//faculty/farabee/BIOBK/14_1.jpg Bacterial Transformation Plasmids Chromosomal DNA Bacterial Cell The uptake of DNA pGLO LAB SUPPLIES FOAM tube holder/float 4 - flip top microtubes Blue- Transforming solution (CaCl2) Yellow- LB nutrient broth Pink- label + Purple- label - 1- colored eraser (to ID your tubes in water bath) 1-pkg yellow innoculating loops 2- Sterile pipettes 4 poured agar plates 1 - LB 2 - LB/amp 1- LB/amp/ara PERMANENT MARKER Cup with crushed ice LABEL TUBES Purple = + pGLO pink = - pGLO Use sterile pipette to add 250μL transformation solution to pGLO + and – tubes Transformationsolution (CaCl2) Get your rack on ICE! INNOCULATE TUBES WITH E. coli BACTERIA Pick one colonyTwirl loop in +pGLO tube Get new loopPick one colonyTwirl loop in –pGLO tube USE SPECIAL GARBAGE BAG FORDISPOSAL OF USED LOOPS EXAMINE pGLO plasmid DNA Use UV light to examine pGLO plasmid vial UV light can be harmful to your eyes!Wear your goggles.Do not shine in eyes. GFP =Green Fluorescent Protein isolated from jellyfish USED AS A GENETIC TOOL http