Hi guys,
Have anyone try to do the synchroizing worms outlined in the worm book? http://www.wormbook.org/chapters/www_strainmaintain/strainmaintain.pdf (Look for protocol 4 and 6)
I have some questions regarding this matter:
= In the protocol, it seems unreal to extract eggs from just 4 worm plates for incubation in 250ml of M9 overnight
= How can we be sure that aspiration too many times won’t loose eggs?
= Where it says in protocol 6: transfer worms to culture inoculated with concentrated E.coli OP50 but does not dictate what concentration and by how much (in protocol 2, it does mention about concentrated E.coli culture, but says that is obtained from spinning down 2 LITRES of overnight E.coli ???)
= Okay, assuming that we get a synchronous population of worms, I want to take this population at L4 stage to do my infection. If I feed the L1 starved worms with E.coli till they grow into L4 (as shown in protocol), how can I obtain a “clean” population of worms to do my infection?
I’ve tried out the protocol once, but did not see any worms hatched even though I did see some eggs after the extraction procedures. I suspect I have lost them through too many aspiration.
Any suggestions?
Thank you very much for your help.