BISC 219/F10: Lab 2 Mutant Hunt
Lab 2: Start Forward Genetics Project: Mutant Hunting
To Do Today (per group)
- Choose one of the six plates of "mutagenized" worms at the front of the room
- Scan the “mutagenized” worms on this plate for visible dumpy mutants
- Transfer 1 putative Dpy mutant hermaphrodite to each of three new plates. Take care not to transfer any other animals or eggs other than your putative mutant.
- Label these 3 identical plates with the hermaphrodite mutant's strain information and your group's information (team color, initials, lab day, date). Use your BLUE Sharpie for labeling these linkage analysis plates.
- Put an elastic around the three linkage determination plates. Place them in your group's plastic box (make sure your box is labeled with your names and lab section on a piece of your team color tape).
- Place your box on the shelf in the incubator for your lab section.
- Incubate your worms at 23°C for 3 days.
3 Days After Lab
- Examine your three plates containing your Dpy worms from the mutant hunt and their F1 progeny. Are all the worms on at least one plate of the Dpy phenotype? If YES, then you are good to go on with the next cross. If you don't have all dumpy mutants in the F1 generation on a plate, you should not use that plate to select worms for the next cross. If none of your 3 plates contains all dumpy mutants, email your instructor ASAP!
- If you have an appropriate plate for continuing: Set up a pair of replicate crosses between 4 Dpy L4 hermaphrodites (should still have the vulval clearing about 1/2 way down the body of the worm) and 3-4 wild type male worms. You will have to search to find the N2 male worms among the hermaphrodites in the wild type worm plates provided in your lab day's box at the back of the room. When you are finished, you should have two identical mating plates with 3-4 L4 hermaphrodites and 3-4 wild type males.
- Label these identical plates with the hermaphrodite mutant's strain information and your group's information including the date. Use your BLUE Sharpie for labeling these linkage analysis/mapping plates.
- Incubate your worms at 23°C until next lab period.
Assignment
Remember to check the Assignment section of the wiki for instructions about the graded assignment due in the next lab and check the Weekly Calendar for other work to accomplish before the next lab.
Links to Labs& Project Info
Series1:
Worm Info
Lab 1: Worm Boot Camp & Sex-Linked or Autosomal Start
Lab 2: Sex-Linked or Autosomal Finale
Series2:
Background: Classical Forward Genetics and Gene Mapping
Lab 2: Mutant Hunt
Lab 3: Linkage Test Part 1
Lab 4: Linkage Test Part 2, Mapping and Complementation
Lab 5: Finish Complementation; Mapping Con't
Lab 6: DNA sequence analysis; Mapping Con't
Lab 7: Complete Mapping: Score
Series3:
Schedule of Reverse Genetics Project
RNAi General Information
Media Recipes
Lab 5: Picking your gene to RNAi
Lab 6: Cloning your gene of interest
Lab 7: Picking your transformant
Lab 8: Plasmid purification and transformation
Lab 9: Induction of bacteria for RNAi
Lab 10: Scoring your worms and RNA purification
Lab 11: RT PCR reactions
