I am evaluating the best disinfection method for in vitro culture of a wild plant. What is the minimum required time for the experiment? Is one month enough?
hi saba
No!
Wild plants are often internal with contamination. Some of which six months after their show.
A simple way to know if your culture are infected is to do a LPGA test.
LPGA: 20g/l Glucose + 7 g/l Peptone + 7g/l Agar , ph 7
put some of your explant (some waste when you do transfer on new medium) an place it in dark, 20-25°C for a week. It will show the majority of possible bacterial infection, even if you can not see them in cultures. For fungi, if you cannot see some after 15 days-1month, I think that your cultures are free.
Sometimes the infection is due to endophytic bacteria. These bacteria become active after a few subcultures, probably when the effect of PGRs decrease. These in-born bacteria in the explants can't be eliminated through ordinary surface sterilization procedures. Under such circumstances, the only solution is use of antibiotics in the medium. We had faced a similar situation with Jatropha curcas. We tested several antibiotics and finally the problem was successfully resolved. Two passages on antibiotic helped us recover our cultures.
@ Tarun Kant: thanks for your explanation, but can you please let me know which antibiotic was good for your experiment and at what concentration?
As i know PPM is not available here in Iran , but it is possible to order it from abroad and as i know ppm is not very effective to reduce internal contamination it will stop bacterial growth outside of explant and airborne contamination in medium but PPM don't remove internal contamination , I have used Cefotaxime and some other antibiotic to prevent or reduce internal or airborne bacterial contamination .if you are interested to have a list of easy to find antibiotics you can contact me .
Ok for the use of antibiotics. But at high concentrations they colud be toxic to
some cell lines
HgCl2 only for bacterial contaminations with moderation: it is very toxic, but often decisive.
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