Pseudomonas aeruginosa Microarray Protocol

互联网2013-09-06

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M. Wolfgang, A. Goodman, M. Gilmore, S. Lory

Department of Microbiology, Harvard Medical School

Important notes before you begin:

· A set of reagents should be purchased for RNA work only. Label these and keep them separate from general lab chemicals.

· Powder free gloves should be worn at all times when working with RNA and RNase-free reagents.

· RNase-free sterile aerosol barrier tiPS should be used for all procedures.

· Store RNA samples at –80oC, and avoid repeated freeze/thaw.

· Treat gloves and work surfaces with RNase Away frequently.

I. Reagents & Materials

The materials section lists recommended reagents and supplies. Many of these may be substituted with an equivalent product, but those listed are known to work well. Not all reagents are specifically listed as RNase-free, however when treated as RNase-free and only used for RNA related procedures, they have worked well.

· 15 mL conical tubes, Corning #430052

· 100% Isopropanol, Mallinckrodt AR #3032

· 1.7mL microfuge tubes, Marsh Bioproducts, #T5050G

· RQ1 Dnase, Promega, #M610A

· RNeasy 50 kit, Qiagen, #74104

· Sea Kem LE Agarose, BioWhittaker Molecular Applications, #50000

· Northern Max-Gly Gel Prep/Running Buffer, Ambion, #8678

· Glyoxal Sample Loading Dye, Ambion, #8551

· 0.24-9.5 Kb RNA ladder, Life Technologies, #15620-016

· RNase-free water (Sterile water for injection), Baxter #2F 7115

· RNase Away, Fisher, 53225-514

· Lysozyme, Sigma #L-7651