BISC 219/2009:RNA interference

These labs were developed with the help of the Silencing Genomes project of the Dolan DNA Learning Center. RNAi General Information Media Recipes Lab 4: Picking your gene to RNAi Lab 5: Plasmid DNA isolation and transformation Lab 6: Induction of RNAi plasmid and C. elegans feeding Lab 7: Single Worm PCR and Agarose Gel Electrophoresis Lab 8: PCR Reaction Cleanup and Sequencing

Schedule of Experiments
{| border="1" ! Lab # !! Dates !! Activity !! Outside lab time ! 4 Set up an overnight culture of your colony !5 Transformation of the isolated plasmid into the HT115(DE) feeding strain Check control and transformation plates for growth - save your transformation plate The night before next lab: Set up an overnight culture of a single colony from your transformation !6 Seed plates and dry for bacterial feeding RNAi Pick 2 L4 hermaphrodite worms of N2 and rrf-3 genotype to 2 RNAi plates for each genotype and make 1 control plate for each genotype as well (6 plates total) Incubate at 23°C until next class !7 Single worm PCR of N2 and RNAi treated worms Agarose gel electrophorsis of PCR products !8 Run sequencing reaction Clean up sequencing reaction with columns
 * 9/29 - 10/5
 * Pick a single bacterial colony containing a plasmid for RNAi
 * The night before next lab:
 * 10/6 - 10/14
 * Plasmid DNA isolation
 * The next day:
 * 10/19 - 10/23
 * Induction of the bacteria to produce RNA
 * 4 days later:
 * 10/26 - 10/30
 * Examine the phenotypes of the fed worms - compare to control N2 worms and worms containing a mutation in the gene you are examining
 * 11/4 - 11/10
 * Clean up PCR reactions with Qiagen Min-Elute kit
 * Analyze your sequencing data