Hello, all friends
I’d like to know your views on the factors that may lead to the death of nematodes.
This death has the following characteristics:
1.Within 3 hours, all nematodes crawled from normal to death.
2.It can be transferred between NGM plate.
3.No obvious contamination of OP50 bacteria and NGM plate was found.
4.There is a cross generational phenomenon.
I’m sure this factor is not temperature. Have you met the same situation?
Any suggestions?
Thank you!
Sounds amazing. The experimental details are unclear though.
Sounds like maybe you made a batch of NGM that kills worms but not OP50(?)
How much can you dilute the “lethal factor” and still cause lethality when added to a “normal” NGM plate?
Is “lethal factor” resistant to high temperature?
By transgenerational effect do you mean that eggs from killed worms fail to hatch?
Hello，
I am convinced that NGM is not a lethal factor, our NGM board is uniformly configured, and OP50 I obtained from another member,and the nematodes raised by him are not abnormal（Consistent OP50 and NGM）.
Cross generational effect does mean that the killed nematode eggs cannot hatch. Even after cleaning, the hatched eggs will suddenly die within 2-3 days of culture.
I tried to culture nematodes with the same strain at 25 ℃, 20 ℃ and 15 ℃, without exception, they could not survive.
However, other members of a bench do not encounter the same problems.
Another strange thing is that the LB I configured will always pollute miscellaneous bacteria without any other treatment after high temperature and high pressure sterilization, while others will not (the same sterilization conditions and almost consistent configuration operation). That I’m not sure if I’m the biggest killer.
Thanks
I can make sure I wear gloves when configuring LB and transferring nematodes, and I will disinfect with 75% alcohol before the experiment.
I tried to put the NGM plate with fatal factors together with the antibiotic free LB plate. There was no colony on the LB plate. I guess it may not be the bacteria themselves that caused the death of nematodes.
Happy New Year 2022！
Check Salt Conc in the solutions like M9. If you are starting from bleach, the time and cracking stuff open needs to be monitored minute by minute. I think.
Thank you
I will check my M9. In the clean operation, I did not use M9, but picked up 5-9 adult with picker and treated with 20ul bleach solution. Generally, after overnight, the larvae will transfer to op50. At this time, there is no obvious abnormality. However, within 2-3 days, they and their offspring will all die in a short period of time, and no obvious signs can be observed before death. I will observe nematodes on a time scale of minutes.
Best wishes
It’s probably either the particular strain you are working with or the media you are using. Swap worms and reagents with your benchmate(s) and see if they can reproduce your results.
Thank you for your advice！
I am now basically sure that factors in my bench environment caused the death of nematodes.
I obtained N2 from laboratory members and cultured it in a 20 ℃ incubator. These N2 survived and could be subcultured.
Then, I asked the laboratory members to help me transfer these N2 to the new OP50 plate with picker, and took 3 pieces and put them in the 20 ℃ incubator; Took three pieces and put them on my bench.
Surprisingly, N2 in 20 ℃ incubator can survive normally. I am currently doing subculture to observe whether their offspring can survive normally.
Most of the N2 placed on my bench died overnight.
I am sure that these N2 are transferred to the same batch of OP50 NGM boards by one person with a picker.
So I think what factors on my bench caused the death of nematodes.
The N2 obtained from laboratory members helped me eliminate the problem of nematodes, and the LB plate without antibiotic activity helped me eliminate the problem of bacteria transmitted between plates.
I am not sure whether the virus plays a role in my bench environment, but it is more likely that some chemical substance or physical factor (excluding the influence of temperature and NGM board) caused the death of nematodes.
It seems that this factor does not kill all N2, a few nematodes survive, and nematodes in starvation or arrest state seem to survive better than others. Interestingly, once these starve or arrest nematodes are transferred to the OP50 NGM plate with picker, they will die quickly.
These are all my current views on this fatal factor. Thank you for your concern and suggestions.