Hello everybody!
I trying to maintain samples of Gelidium elengans for protoplast isolation in sterile Petri dishes containing 100 ml of PES medium at 20°C and medium light intensity (2 fluorescents lamps). However, 1 or 2 days after placing the samples in the medium, most of the apical tips of the main axes and the branches became bleached. Then, I changed the temperature to 15°C and I put the samples in low light intensity, but this only slowed a litle the bleaching.The medium and basal parts of the main axes didn't change color. I was wondering what environmental factor would be affecting my samples.
Many thanks in advance
Dear Jose,
There could be many factors for the bleaching of seaweed in culture like the duration and intensity of light or temperature.Could it be due to some bacterial growth?.
Why don't you culture few fresh samples under a range of the above mentioned factors or even in slightly modified medium? (if possible)
Good luck!
Thanks Jackson! I'm trying to modify the concentration of N in the medium as I see in some papers that bleaching could be caused by excess of light or N depletion.
Thanks again
Dear Peltroche, Bleaching of Gelidium bits in invitro conditions maybe due to any one this these
1) Lack of proper osmoticum of the medium- check the pH and osmoticum similar to the tissue
2) Variation in salinity - adjust the salinity of the medium to the ambient levels from where it was collected or accilmatized.
3) Maintain a photoperiod of 10/14 hr of L / D. and
4) The Most important is temp from light source- Adjust the proximity of lamp to the Petri plates or use a water filter between the two
Wish you god luck.
Dear José,
I agree with previous comments about light intensity + N. Also:
1. What volume are the Petri dishes? Are they being agitated? If they are shallow and not stirred than nutrients may be depleted near fronds.
2. Can you measure light intensity with a quantum sensor (micro moles/ s/ m)? 2 fluo lamps may be too much if very close. May also be problematic for temp.
Check if the fronds are growing at all, or remain the same length. Also check under microscope for epiphytic microbes.
Best wishes
Dear Ricardo,
I have reduce the light intensity to a 2-3 umol. Also I have increased in 25% the NaNO3 concentration in PES medium and it seems that the bleaching stopped. I have new Gelidium samples and I put them in this PES modified medium and, until now, there is no bleaching in them. Also, I think there is some bacterial contamination in the bleached parts (I saw this in the microscope).
Many thanks
Olá José,
That is great to hear! What species are you working with? In my experience smaller species and larger ones behaved very differently in the lab. Larger spp - like G. robustum and G. corneum - grew much better from prostrate branches. And they really liked agitation.
Again, best of luck with your project.
Many thanks Ricardo! I am working with Gelidium elengans! The main idea is isolated protoplast from this species in order to stablish cell suspension cultures :) Many thanks for your help again!
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