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By avadakedavra
Posts:  90
Joined:  Thu May 18, 2017 2:06 pm
#300225
I have some VFT seeds and sow in vitro.

Some of them have great succeed.

but others contaminated.

It looks the pollutions are from inside of seed, I guess.

Image.

What can I do?

May I sterilize the seeds again and sow ?

Or may I give up TC culture and sow them on the other medium like moss ?

success seeds are all succeed (same packet from my friend )

high contaminated seeds , more than half of them are polluted and none of them germinated. These are imported seeds.

I cannot decide how to care.



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By xr280xr
Posts:  2806
Joined:  Wed Jun 22, 2011 3:29 pm
#300301
If the seeds' survival is very important to you, I would probably try to gently re-sterilize them or even just rinse them really well and put them in sphagnum moss. If they haven't germinated you could always try full sterilization again and put them back in TC, but it seems like they'd have a lower success rate. You may want to re-evaluate your procedures and look for possible sources of contamination slipping by too.
By nicszabo
Posts:  16
Joined:  Sat Feb 21, 2015 6:22 pm
#301082
"You may want to re-evaluate your procedures and look for possible sources of contamination slipping by too." xr280xr

One very likely source could be your autoclaved water used for rinsing. Make sure to measure the water you dispense into containers and follow an autoclave times chart.

Here is an example: https://phytotechlab.com/media/document ... tMedia.pdf

One way to test this out is to dip your sterile forceps, scalpel, etc. into the water used for rinsing and place it onto media. Wait 3-5 days and evaluate.
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By xr280xr
Posts:  2806
Joined:  Wed Jun 22, 2011 3:29 pm
#301098
nicszabo wrote: One way to test this out is to dip your sterile forceps, scalpel, etc. into the water used for rinsing and place it onto media. Wait 3-5 days and evaluate.
Media without PPM or similar biocide.
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By avadakedavra
Posts:  90
Joined:  Thu May 18, 2017 2:06 pm
#301461
xr280xr wrote:
nicszabo wrote: One way to test this out is to dip your sterile forceps, scalpel, etc. into the water used for rinsing and place it onto media. Wait 3-5 days and evaluate.
Media without PPM or similar biocide.
thank you for your advice
I've tried and a few of them are contaminated again.
I'll try moss.


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User avatar
By avadakedavra
Posts:  90
Joined:  Thu May 18, 2017 2:06 pm
#301463
nicszabo wrote:"You may want to re-evaluate your procedures and look for possible sources of contamination slipping by too." xr280xr

One very likely source could be your autoclaved water used for rinsing. Make sure to measure the water you dispense into containers and follow an autoclave times chart.

Here is an example: https://phytotechlab.com/media/document ... tMedia.pdf

One way to test this out is to dip your sterile forceps, scalpel, etc. into the water used for rinsing and place it onto media. Wait 3-5 days and evaluate.
thank you for your advice
contamination and germination occurred like 'all or nothing '

I'll check my method of sterilization again.


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