Hello, colleagues.

I had this problem in my in vitro cultures of Cnicus benedictus, which I derived from immature embryos. After several months of vegetatively multiplying the plants to get a lot of clones from each genotype, which I further want to use for polyploidization with oryzalin, I have experienced endogenous bacterial contamination. The senior doctor, who is a specialist in in vitro culture in our department, says that some embryo cells may already have had endogenous bacterial contamination, and now that I've cut the plant tissue while propagating, we've come across the cells, and the contamination has gone to the media. Does that seem likely?

I use 100 microliters per 100 milliliters of PPM (Plant Preservative Mixture) and, I'm recommended to increase this concentration three times. If that doesn't help, I'd have to use the media using antibiotics, but I'd like to avoid that because it would take me at least a week of passage and another week of passage back to classic MS after a week.

What would you recommend that I don't have a similar problem in the future? Alternatively, what are your pointers for handling the current sticky situation?

Many thanks in advance for the suggestions and good day and good health.