Lab 2: Start Forward Genetics Project 2: Testing a Mutant for True Breeding
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 and determine their phenotype. Make sure you write this down.
Transfer 1 putative mutant hermaphrodite to each of two new plates. Take care not to transfer any other animals or eggs other than your putative mutant.
Label these 2 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. Labels belong on the side of the plate with the media in it. Tape makes it hard to see the worms on later days. DO NOT write on the lids - they fall off and you will not know what cover goes to what bottom!
Put an elastic around the two phenotype 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 two plates containing your mutant worms 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 neither of your 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 wild type 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.