Isolation of microalgae
Posted 22 August 2011 - 07:22 AM
at the moment I am trying to isolate some microlgal species from field samples. The two most common techniques I found were streaking on agar (with medium) and serial dilution method.
Textbooks and research papers make these methods sound do easy and straight-forward, yet for me, they are anything but.
To start with, inoculation on agar gave me too much bacterial and fungal growth, with no (or really few) algal colonies. I incorporated Pen/Strep and recently, Amphotericin B, in the agar, and growth is still poor.
I carried out the serial dilution method in 96-well plates. Even here, many wells had extensive fungal growth. I will try to incorporate Amphotericin B in these as well, although I am a bit wary that it may affect the microalgae.
I am working aseptically, and the fungal source is most probably from the samples themselves, since each sample has a characteristic fungal growth.
I was just wondering if anyone else had similar problems with such contaminations?
In addition, some protocols state that when preparing agar plates with medium (e.g. BG-11), you need to autoclave the salts separately from the agar. I have not done this yet, although I am planning to prepare a batch like this. Do you think it really makes a difference?
I would really appreciate your feedback.
Posted 22 August 2011 - 10:11 PM
Posted 05 September 2011 - 05:07 AM
The problem is that I am not getting any algal colonies at all - so I cannot re-seed them.
I also prepared plates with agar and salts autoclaved separately, and still, no colonies.
Posted 05 September 2011 - 07:01 AM
Posted 05 September 2011 - 07:29 AM
Could be that the Pen/Strep and amphotericin B themselves are inhibiting algal growth, I guess.
I am quite at a loss; as I thought this process would be much more straight forward. I may have to skip the isolation process and get hold of pure cultures.
Posted 05 September 2011 - 09:41 AM
Posted 12 September 2011 - 05:23 AM
I use distilled water for preparation of agar and media. These are always autoclaved before use.
To be honest lately I have been noticing some algal colonies amidst the fungal and bacterial ones. I am trying to re-streak the few colonies I have. In the meantime, I'll keep my fingers crossed.
I am also preparing the same plates without Pen/Strep, as I read that Streptomycin may cause chloroplast bleaching in some green algae.