Summary:
We recently published a piece on the importance of temperature in C.elegans which got the whole InVivio Biosystems team thinking about C. elegans’ development, and how it can best be compared. As a result, we thought it would be fun to take pictures of C. elegans in the three most commonly used temperatures to emphasize how warmer temperatures cause C. elegans to age faster. This protocol will walk through the process of this experiment, to read more about how temperatures affect C. elegans, check out our recent blog.
Day 0:
The worm population is synchronized through bleaching, this eliminates differences based on the worm’s age at the start of the experiment.
Image 1. C. elegans bleaching protocol (generated in BioRender).
Day 1:
Morning - Worms are plated on differing plates for each of the temperatures we are looking at (1x, 2x, 3x). Allow the plates to dry completely before moving to the next step..
10:30 - First round of pictures are taken of each plate (each temperature) at 3x and 4.5x. At least six images for each plate are captured.
Afternoon - Repeat the procedure from morning plating (worms on differing plates for each temperature and allow to dry).
16:30 - Second round of pictures are taken of each plate (each temperature) at 3x and 4.5x. At least twelve images.
Day 2:
10:30 - Pictures are taken of each plate (each temperature) at 3x and 4.5x. At least twelve images should be captured.
16:30 - Pictures are taken of each plate (each temperature) at 3x and 4.5x. At least twelve images.
Repeat for 5 days (96 Hours)
Stop taking pictures when plates contain adults [see table below].
- 16C stopped taking pictures at 138 hours
- 20C stopped taking pictures at 78 hours
- 24C stopped taking pictures at 72 hours
| Wed | Thur | Fri | Sat | Sun | ||||
|---|---|---|---|---|---|---|---|---|
| 2/16 | 2/17 | 2/18 | 2/19 | 2/20 | ||||
| Worms plated at: | time to take images: | Temp: | D1 | D2 | D3 | D4 | D5 | Plate labeled: |
| AM 2/16 | 10:30 AM
(1030) |
16C | 0 hr | 24 hr | 48 hr | 72 hr | 96 hr | N2 2/16 AM 16C |
| 20C | N2 2/16 AM 20C | |||||||
| 25C (24C) | N2 2/16 AM 25C | |||||||
| 4:30 PM
(1630) |
16C | 6 hr | 30 hr | 52 hr | 78 hr | 102 hr | N2 2/16 AM 16C | |
| 20C | N2 2/16 AM 20C | |||||||
| 25C (24C) | N2 2/16 AM 25C | |||||||
| PM 2/16 | 10:30 AM
(1030) |
16C | X | 18 hr | 42 hr | 66 hr | 90 hr | N2 2/16 PM 16C |
| 20C | N2 2/16 PM 20C | |||||||
| 25C (24C) | N2 2/16 PM 25C | |||||||
| 4:30 PM
(1630) |
16C | 0 hr | 24 hr | 48 hr | 72 hr | 96 hr | N2 2/16 PM 16C | |
| 20C | N2 2/16 PM 20C | |||||||
| 25C (24C) | N2 2/16 PM 25C |
Conclusion:
This experiment was a fast and simple way to show the significant impact that temperature can have on C. elegans’ development - worms in the 24C condition progressed to egg laying stage in nearly half the time as the worms in the 16C condition [Image 2].
Image 2. Development of C. elegans in different temperatures over several days.
Thus, this experiment emphasizes how important it is to carefully consider your husbandry before embarking on a study. While all the temperatures we used in this experiment are regularly used in C. elegans husbandry, the worms vary greatly and this could impact your study’s results.
About the Authors
Thomas is a Technician at Invivo Biosystems. He graduated from University of Oregon in 2019 where he earned a Post-Baccalaureate Certificate in Fisheries Management. He has worked as a scientist for NOAA NMFS SWFSC, surveying presence and abundance of juvenile pelagic fish species along the west coast of the US. He joined the Invivo Biosystems team in 2022. He swims and plays board games in his free time. Also, he enjoys giving presentations and teaching, outreach is a passion of his.