Help to sterilize unknown seed - (Dec/27/2013 )
I have to do tissue culture of the seed (given an unknown seed, name as seed x)
i culture it 3 times already but damaged du to contamination, what can i do to avoid the contamination and make the seed germinate?
first technique: put under running tap water - 15 min
under laminar flow: put with 70% ethanol for 3 min
50% clorox-10 min
2nd sterilization 95 % ethanol- 1/2 min
25% clorox- 5min
results: contaminated
2nd technique: seed soaked overnight
run under running tapwater- 30 min
70% ethanol- 1 1/2 min
50% clorox with tween 20- 10 min
result: contaminated
the seed picture- in the attachment
the methods you use seem ok.
2 reasons that might be causing the problems:
- your technique working in the hood is not good enough (which is a known problem when working with seeds with people that do not do this on a daily base, I am not sure whether you are a student or used to this?)
- contamination in the seeds themself, under the "skin".
There are many protocols to sterilise seeds, so you might want to look for an alternative, but yours seem (at first sight) ok.
i used to do sterilization, but this seed is given as unknown, i dont know the exact protocol to use. yes, i thought i saw some fungus inside the seed (in the second picture of the attachment)
If there is a fungus inside it, you will need to do some other stuff. But I dont know the seed, could it be possible to remove the outer protective layer and then sterilize it and let it germinate? If there is an external layer that is contaminated, this might be a option.
Other options: just grow the plants with the intact seed, without sterilisating it and then work from the plant material....
i used to do sterilization, but this seed is given as unknown, i dont know the exact protocol to use. yes, i thought i saw some fungus inside the seed (in the second picture of the attachment)
I am typically use PPM product to sterilize the untamed explant. it is very powerful.
You can check this one out
http://www.plantcelltechnology.com/plant-preservative-mixture-ppm/
Guys - please use correct terminology. You are not "sterilizing" anything with those protocols.