Download RNA Viruses: A Practical Approach by Alan J. Cann PDF

By Alan J. Cann

This booklet presents a extensive remedy of the foundations and perform of RNA virus learn to insure the widest attainable viewers. it really is excellent for laboratories occupied with virus tradition, together with either educational and commercial examine teams and public healthiness laboratories.

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The pellet should be resuspended in 1/10th the original volume of culture fluid. 2 g of Nad per 100 ml of culture fluid with constant stirring at 4°C, followed by the addition of 6 g of polyethylene glycol 6000 per 100 ml. Stirring should be continued at 4°C for a minimum of 4 h, and the precipitate concentrated by centrifugation at 2000 g for 2 h. The pellet should be resuspended in l/50th to l/100th the original volume of culture fluid (or an appropriate buffer). The latter A. J. EASTON ET AL.

1 Site-directed mutagenesis of infectious clones of RNA virus genomes The in vitro techniques that facilitate the mutagenesis of RNA viruses rely on site-directed mutagenesis of cloned cDNA. This is performed using standard recombinant DNA methodology with oligonucleotide-directed mutagenesis of an infectious cDNA clone of the virus genome. Often the mutagenesis is facilitated by PCR, in which one of the primers encodes the desired mutation as a mismatch to the wild-type sequence. If the mutation is close to a naturally occurring restriction enzyme site within the cDNA sequence, then the replacement of the wild-type sequence with the mutation is relatively simple.

0 with sodium hydroxide. 22 um filter or autoclaving. Formaldehyde 10 mg/ml ethidium bromide in water. Store in the dark. 1% w/v) Ultraviolet transilluminator Method 1. 2. 3. 4. 5. 6. 7. 8. 16 Dissolve the agarose by boiling in the appropriate volume of 1 x MOPS buffer. 22 M final concentration. Pour into a gel-former and allow to set in a fume hood or a closed box. 1 ul ethidium bromide. Denature by heating to 68°C for 10-15 min. Cool and load on to the gel. Load Bromophenol Blue marker dye into one or more wells.

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