Recombinant DNA PPT _3 2.24.11BNS

Recombinant DNA PPT _3 2.24.11BNS - fragments Supercoiled...

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LOADING YOUR GEL Preparing your samples A. Add 2µl of tracking dye B. Centrifuge to spin down sample Loading the gel    - TA will do a DEMO A. Submerge gel in 1X TAE buffer in gel rig B. Slowly remove comb - pull straight up C. Load your sample into the well D. Connect electrodes --  DNA will move from  negative (black) to positive (red) E. Run gel at 120 volts Video 
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POURING AN AGAROSE GEL 1. Making the agarose gel: A. Weigh out the necessary amount of agarose B. Add two pumps of TAE buffer C. Microwave for one minute D. Swirl (CAREFULLY!) E. Microwave again to complete melt agarose 1. Pour gel into the tray and insert the comb.
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Agarose Gel Electrophoresis + - Add DNA samples into wells Apply electric charge across gel Since DNA has a negative charge, itwill run towards positive electrode
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AGAROSE GEL ELECTROPHORESIS SIZE SEPARATION USING A GELATINOUS MATRIX + - Small DNA fragments Large DNA
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Unformatted text preview: fragments Supercoiled DNA Linear DNA Open circular Molecular Weight Markers DNA fragments of known sizes Compare your DNA bands to determine approximate size ***This ladder is accurate only if DNA is linear. Circular and supercoiled DNA run at different rates and will not correspond with the DNA ladder Ethidium Bromide Staining of DNA Ethidium Bromide Intercalates into groves of DNA. It fluoresces under UV light. CAUTION: EtBr is a carcinogen. Capturing your Gel Image Your TA may have to do this for you depending on time Transilluminator-UV light source Camera HOW TO LABEL AN AGAROSE GEL Lane Assignments: 1.MW 2.Uncut 3.PstI plasmid #1 4.EcoRI 5.Double digest EcoRI & PstI 1 2 3 4 5 Name MW Labels Mapping Tutorial Go through the Tutorial on Restriction Mapping Questions #5 and #6 on datasheets NEXT LAB Restriction Mapping Quiz over module...
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Recombinant DNA PPT _3 2.24.11BNS - fragments Supercoiled...

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