I have such a problem that in a single colony there are 3 different bacteria. How can I easily purify them? Do you have any suggestions? Thank you.
Dear Ebru,
it often helps to cultivate a mixed colony in liquid culture for a reasonable number of generations. Reasonable means: a sufficient number of cells per ml but less than allowing the bacteria to embedd themselves in too much EPS jelly. Very often the bacteria have then formed small aggregates of the same type that, after streaking again on solid medium, develop as pure cultures. If you are especially lucky, your bacteria will even stop to secrete EPS in liquid culture. Just check in the microscope.
Good luck. - Johannes.
Dear Ebru,
prepare further dilutions.
Add chemical reduce EPS production.
Dissoleve some of the EPS and streak it again on solid medium.
Hi,
Make dilutions and plate it. you will get single colony from which you can purify your isolate.
Dear Professor Wöstemeyer, Dear Proffesor Dave, Dear Dr. Viswanathan firstly thank you very much for your answers. I used very different techniques. I inoculated 1 uL of liquid culture to solid medium. I made dilutions and pour plate techniques in three different times. I used single colony in every streak plate method. After doing every step, I checked gram stain reactions. Once, I used 1% of Triton X-100 in dilutions also. But this methods didn't work in order to purify the different bacteria. But then I thought to look petri for colony morphology on microscope and then I saw the different bacteria on that colonies because of EPS. I can also give pictures of them. So at the end I saw some colonies purified some colonies were the same that they are not purified. I will use the purified colony but I also wonder to use the other colonies. So which chemical is the best in order to reduce the EPS production easily?
Thank you very much again,
Yours sincerely,
Ebru
Hi,
What is the media you are using for culturing this organism? Some media may contain sugar which may enhance the production of EPS. Try to streak the bacterial colony in minimal mineral media without adding sugar.
Thank you Ebru,
I am not convinced for all of the pictures that you show that all of these examples are really mixed cultures. You could, with some misfortune, just have plates or microphotographies of colonies with a somewhat deviating morphology, at least in some of your pictures. Maybe most of your isolates are purer than you think. You could check individual colonies from your isolation procedures by one or the other fingerprinting technique, mybe ERIC-PCR or similar. Just give your experiments a chance. In principle you did everything the right way.
Good luck - Johannes.
Dear Professor Wöstemeyer,
Thank you very much. I am going to select the colonies for genetic analyses but now I have universal primers after that I will also use ERIC-PCR. Thank you for your support and suggestions.
- Badges
- Science method
- Similar topics
- Biological Science
- Ecology