Hi Bart and all,

In general answer is YES. I am working on tree species form temperate hemisphere so most of them are dormant (deeply dormant) with different types of dormancy, and on the basis my experience I could say that they stain red if there are alive (you could see some results i.e. in my last publication in Tree physiology titled: Changes in genomic 5-methylcytosine level mirror the response of orthodox (Acer platanoides L.) and recalcitrant (Acer pseudoplatanus L.) seeds to severe desiccation (I have attached PDF file) both investigated Acer species are dormant. We are also finishing our paper on storage and desiccation Fagus sylvatica (also dormant seeds) where the seeds were stained red in TTC. My former head of laboratory prof. Tylkowski made a lot of such studies unfortunately many of them are not available in ISI Web of Knowledge and were published in magazine Arboretum Kórnickie (luckily in English). There are also reported other works which generally show that TTC could be applied. However as you can observe in our manuscript the results obtained by TTC test are generally higher than by germination test (even conducted in laboratory) it is closer to the results obtained during in vitro regrowth of embryonic axes, and it is also rule which I observe more often.

In my opinion TTC test could be applied only in the case of pure science as indicator if embryos are physiological viable or not, but it does not have any practical aspects and should not be used in this purposes i.e. during trading of seeds between companies. So in my opinion TTC test should not be used by ISTA to assess seed viability as it is not reliable marker of seed viability and shows much higher results than true ability of seeds to produce seedlings. At least this situation is very common in the case of dormant seeds from trees/shrubs species in which I specialize.

Best regards,

Marcin

When dehydrogenase is there, staining will certainly take place. That is why dormant seeds also take stain and are reported viable.

This is one reason why viability percentage shown by TTC test is in most cases more than actual germination in germination test.

I strongly agree with P. R. Kumar. The test TTC is used for seed viability determination - if the plant tissue is in a metabolic activity, tissue turn coloured in dark red. I have no personal experience in TTC vs. dormant seed. But I think that tissue of dormant embryo "breathes less", and colour will be more light (red) in dormant tissue after TTC treatment, than colour in non-dormant tissue (comparison between one plant species).

We know that reduction of water-soluble tetrazolium salt like TTC by metabolically active eukaryotic and prokaryotic cells leads to precipitation of colored formazans. The reaction has been applied in a variety of biological assays including subcellular localization of oxidoreductases. It has also been used to test viability in seeds and to estimate cell culture growth. Reduction of TTC in isolated cells, tissues and seeds is regarded as an indicator of “cell redox activity”. The reaction is attributed mainly to mitochondrial enzymes and electron carriers.

Dr. Panis, I think that you have a very important question, we do not know what happens in the seeds or embryos that have some type of dormancy so deep that it is able to avoid the activity of these enzymes keeping them at zero, which consequently makes the TTC test ineffective.

We also have another question; evidence exists that the TTC reduction in mammalian cells is also catalyzed by a number of nonmitochondrial enzymes but we do not know what happens in the seeds or embryos. In particular, it has not been demonstrated whether the reaction takes place in mitochondria or whether it is catalyzed by nonmitochondrial-reducing species, away from mitochondria.

Regards from CNRG,

Banana Guy!

Hi Bart and all,

In general answer is YES. I am working on tree species form temperate hemisphere so most of them are dormant (deeply dormant) with different types of dormancy, and on the basis my experience I could say that they stain red if there are alive (you could see some results i.e. in my last publication in Tree physiology titled: Changes in genomic 5-methylcytosine level mirror the response of orthodox (Acer platanoides L.) and recalcitrant (Acer pseudoplatanus L.) seeds to severe desiccation (I have attached PDF file) both investigated Acer species are dormant. We are also finishing our paper on storage and desiccation Fagus sylvatica (also dormant seeds) where the seeds were stained red in TTC. My former head of laboratory prof. Tylkowski made a lot of such studies unfortunately many of them are not available in ISI Web of Knowledge and were published in magazine Arboretum Kórnickie (luckily in English). There are also reported other works which generally show that TTC could be applied. However as you can observe in our manuscript the results obtained by TTC test are generally higher than by germination test (even conducted in laboratory) it is closer to the results obtained during in vitro regrowth of embryonic axes, and it is also rule which I observe more often.

In my opinion TTC test could be applied only in the case of pure science as indicator if embryos are physiological viable or not, but it does not have any practical aspects and should not be used in this purposes i.e. during trading of seeds between companies. So in my opinion TTC test should not be used by ISTA to assess seed viability as it is not reliable marker of seed viability and shows much higher results than true ability of seeds to produce seedlings. At least this situation is very common in the case of dormant seeds from trees/shrubs species in which I specialize.

Best regards,

Marcin

Yes a dormant embryo will stain red when treated with TTZ if and only if it the embryo is dead.

The mechanism of TTZ test is based on the fact that all living tissues must respire, thus capable of reducing a colourless TTZ in to a red coloured compound called Formazan. The change in colour from colourless to red is enhanced by H+ transfer reactions catalysed by the dehydrogenase enzyme found only in living tissues. The Formazan being non-diffusible stains the living tissue red. Therefore as far as the embryo is still alive it will stain red when incubated in TTZ .

see attachment for reaction mechanism

sorry, i just discovered the first statament of my answer had a mistake. The intended answer is '' Yes a dormant embryo will stain red when treated with TTZ if and only if the embryo is not dead''.

Dormancy does not affect TTC test results. Viability (% ) can be assessed by TTC test. It gives no surmise of dormancy. Laboratory germination test has to be performed and the difference between TTC and laboratory germination test can be taken as an indicator of level of dormancy in any seed lot.

Agree with @Marcin Michalak.

This link of article may be helpful

Article Salt Induces Features of a Dormancy-Like State in Seeds of E...

Regards

this link is useful

Article Determination of seed viability of eight wild Saudi Arabian ...

regards

Agree with Dr. Kumar. Dormancy is such a complex process that it can not be solved by a simple TTC test.