By M. Demerec (Ed.)
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A accomplished compendium of state-of-the-art protocols for the iteration of molecular range. defined in step by step aspect to make sure experimental good fortune, those protocols contain effectively reproducible equipment for random mutagenesis of whole genes or segments of genes, for homologous and nonhomologus recombination, and for developing in vivo libraries in micro organism and yeast.
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6. Use the chip within 5 min of preparation to prevent evaporation. Cover the chip if it will be left standing for any length of time 7. Launch the 2100 Expert software. 8. Place the chip into the Bioanalyzer and close the lid. A chip symbol will appear on the screen. 9. Click the Electrophoresis button, click Assay, click RNA, and select Eukaryote Total RNA Nano from the menu. 02/Braun/13-40 23 12/20/06, 12:54 PM 24 Schinke-Braun and Couget 10. If necessary, edit the number of samples using the menu on the lower left.
All seven peaks should be well resolved. 3. To review the data for a specific sample, select the sample name and highlight the Results subtab. Major features of an electropherogram for a successful eukaryotic total RNA run are sharp and well-resolved 18S and 28S ribosomal RNA peaks. There should be no or only a small peak for the 5S ribosomal RNA. The baseline should be flat. Smaller additional peaks and a hump in the baseline are indicative of degradation (see Note 8). 4. Microarray gene expression experiments should not be performed with poor-quality total RNA samples.
2 mL, so that it fits snugly under the silver stopper. Press down slowly and steadily on the plunger when priming. 5. Wait exactly 30 s. 6. Squeeze the silver stopper to release the plunger. 7 mL within 1 to 2 s. If it does not prime well, check that the gasket is clean. Change the gasket if the problem persists. 7. Pull the plunger up the rest of the way to 1 mL. Be careful to avoid a negative pressure vacuum, which can lead to bubbles. 8. Lift the priming station lid. Hold the chip in place with one hand and lift the lid with the other to prevent dislodging of the chip from the priming station.