Thanks so much for answering my previous questions. I’m starting my science fair project soon and have some more questions.
- How often do I need to wash the worms? What about synchronizing them? The person I asked said that she only washed and synchronized them once (at the beginning of her project).
- Any tips/suggestions on how to transfer worms from plate to plate?
- I was told to transfer them every three days using a sterilized pick and to make the pick slightly sticky by touching it to the agar first. Once I get the worm on the pick, how do I transfer it to the new plate? Do I let it crawl off?
- Can the light at the bottom of the microscope kill/harm the worms?
- What temperature are the worms supposed to be stored at?
1) How often do I need to wash the worms? What about synchronizing them? The person I asked said that she only washed and synchronized them once (at the beginning of her project).
When you are growing the worms and for many procedures you do with them, it is not necessary to wash them. Washing them becomes important only when materials from their environment (in particular the bacteria) are considered likely to disrupt an experiment, for example if they were likely to metabolize a drug you’re giving the worms. There may be some confusion here between “washing the worms” (to remove material normally present with the worms immediately prior to an experiment) and “cleaning a strain”. You “clean a strain” to remove inappropriate strains of bacteria, yeast, or mold that might have contaminated the culture. This is done to make sure that you know what the culture conditions are and that they are identical for your experimental and control populations. Normally you would clean a strain when you receive it any then avoid (and monitor for) any future contamination of the strain, but not clean it again. Washing, if necessary, must be done each time you perform the experiment. You can read up on cleaning strains in this Wormbook chapter.
There are suggestions and links for how to synchronize worms in this thread.
[i]2) Any tips/suggestions on how to transfer worms from plate to plate?
- I was told to transfer them every three days using a sterilized pick and to make the pick slightly sticky by touching it to the agar first. Once I get the worm on the pick, how do I transfer it to the new plate? Do I let it crawl off?[/i]
There is information on transferring worms in this Wormbook chapter. When making your worm pick (referred to in the linked Wormbook chapter as a ‘worm picker’), you can flatten the end of it (roughly 0.5-1 mm in length) using a pair of smooth-surfaced (not toothed) pliers, rather than a hammer as suggested (use 1 - 1.25" of platinum wire, melted into the end of a 5.75" Pasteur pipet). If there are sharp corners or edges, you can try to file them away as suggested, or you can sometimes cut them off with a sharp razor blade. Sustained heating of the sharp corner can also dull it somewhat. Different people have differing preferences for the shape of the finished pick, but I’d start with the wire in an arc resembling ~90 degrees of the circumference of a circle from the end of the pipet (from ~level to ~straight down), ending with a 90-degree corner to the flattened surface (which will then be level).
It is possible to use the pick like a scoop to pick up worms, but it is difficult to do this without damaging the surface of the agar, especially using a newly-formed pick and with limited practice; moreover, the worms will tend to be reluctant to crawl off of the pick. The best approach is usually to get a blob of bacteria on the underside of the pick by pulling the edge of the flattened surface along the surface of the bacterial lawn (note that the edge of the lawn is usually the thickest part, which is good for this, and drier plates yield larger masses of stickier bacteria; plates seeded with bacteria only 1-2 days previously are less dry and can give smaller, relatively less sticky blobs of bacteria). You can then press this blob gently down onto the top of a worm or swipe it at the side of a worm, and then gently release the worm onto a new plate by reversing the procedure. Both picking worms up and releasing them can be easier from the middle of the bacterial lawn than from its edges or from the bare agar. If it is relevant to your experiment, be aware that your blob of bacteria can simultaneously transfer eggs or young larvae that you might not notice if you don’t carefully check for them; L1 larvae in particular can be hard to spot until they start moving again. Depending on what bacterial strain you are using, with practice you can easily transfer 100 worms on your pick at one time.
3) Can the light at the bottom of the microscope kill/harm the worms?
Too much light can harm worms, especially embryos, although I don’t know if this occurs at the sort of intensities used in a dissecting microscope. Too much heat can definitely harm worms, especially over sustained periods. A lot depends on what sort of scope you are using; if it has a bright bulb directly within the base of the microscope (as opposed to outside and shining into a mirror), the base of the scope can get quite warm.
4) What temperature are the worms supposed to be stored at?
Wild-type strains can be cultured reliably at temperatures from about 12.5 degrees Celsius to something between 26 and 30 degrees Celsius. 10 degrees Celsius is supposed to cause developmental arrest, and 30 degrees will eventually cause sterility. The standard temperature is 20 degrees, and most experiments are done using animals cultured at this temperature. Note that developmental time is dependent on temperature: at 15 degrees, the generation time will be about 5 days, at 20 degrees about 3-4 days, and at 25 degrees about 3 days. Animals are more stressed at higher temperatures. If you are keeping them with you at room temperature rather than in a separate room or incubator, 72 degrees Fahrenheit (about 22 degrees Celsius) can be a good compromise for your comfort that shouldn’t cause problems for most healthy strains.