Secret Project.........

Very interesting. I wish I had thought of something like this earlier. 
 
@geonerd The plant might lose the polyaploid cells later, but this is only if there are normal diploid cells in the plant as well. Seeing as tetraploid should have 4 of each chromosome (correct me if I'm wrong), it would take a lot more effort to reproduce the DNA for cell division. As such, if there are enough normal cells, the polyaploid will get bred out eventually. 
 
Nigel, THANK YOU for doing this. I think you just inspired a project for my molecular and cellular biology class next year. After all, every experiment needs to have multiple trials to ensure it's valid. :clap:
 On another note, you're SURE that TSMB was a good idea for this? That the pods aren't already dangerous to the sanity of whoever eats them? I gotta see what happens if this works.
 
And on the final note, seeing as a karyotype is basically gene staining, a coloring book is exactly what needs to be done here....
 
Edit: Peter pepper that's extra spicy and is a foot long. That;s what you should have aimed for.
 
cruzzfish - most likely anyone claiming a foot long Peter Pepper is a liar or Mr. Ed.
 
Nigel - what was your field of study for your doctorate?
 
cruzzfish said:
Very interesting. I wish I had thought of something like this earlier. 
 
@geonerd The plant might lose the polyaploid cells later, but this is only if there are normal diploid cells in the plant as well. Seeing as tetraploid should have 4 of each chromosome (correct me if I'm wrong), it would take a lot more effort to reproduce the DNA for cell division. As such, if there are enough normal cells, the polyaploid will get bred out eventually. 
 
Nigel, THANK YOU for doing this. I think you just inspired a project for my molecular and cellular biology class next year. After all, every experiment needs to have multiple trials to ensure it's valid. :clap:
 On another note, you're SURE that TSMB was a good idea for this? That the pods aren't already dangerous to the sanity of whoever eats them? I gotta see what happens if this works.
 
And on the final note, seeing as a karyotype is basically gene staining, a coloring book is exactly what needs to be done here....
 
Edit: Peter pepper that's extra spicy and is a foot long. That;s what you should have aimed for.
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I`m 100% positive that TSMB was the right choice........Bwah ha ha. 
 
Doing multicolor FISH would be very cool, but perhaps beyond my means at this point. I`ll settle for a yes or no answer!
 
Hmmm, foot long, hot Peter Peppers. You should go for it! 
cone9 said:
cruzzfish - most likely anyone claiming a foot long Peter Pepper is a liar or Mr. Ed.
 
Nigel - what was your field of study for your doctorate?
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Hahahahah.
 
My PhD was in Biochemistry, postdocs in Cell Biology, Molecular Biology, Genetics and Neurobiology. 
 
Here`s a photo of one of the seedlings from the 0.1% colchicine. Weird does`t do it justice! I now have 6 that look exactly the same from the 0.1% seeds, 4 or 5 that look the same from the 0.05% seeds and a couple of normal plus a couple of these from the 0.025% seeds. My first thought is that the time of treatment was too long, at 72 hours. These weird seedlings don`t seem to grow much and do not put out a tap root very far. 
 
 
Awesome, Nigel! 
 
*I'm a little seedling short and stout!*
 
I can't wait til these guys set pods! Do you have any (High Performance Liquid Chromatography) HPLC equipment to do capsaicin analysis? If you did an assay across all your plants with an N=3 or N=5, you could get dope results on the efficacy of colchcine on increasing capsaicin in pods. Plant size, growth rate, pod production and size would be easy to quantify but I doubt you'd want to taste test the capsaicin content of these suckers. I'm really excited to see these results. 
 
Also, do you think measuring optical density of a capsaicin solution would work? You could definitely extract the capsaicin with methanol or hexane and then measure the OD and backtrack to calculate the concentration with the Beer-Lambert Law. If you don't have HPLC and you or your wife got an A in orgo, it wouldn't be too difficult. Just time consuming.
 
SciurusDoomus said:
Awesome, Nigel! 
 
*I'm a little seedling short and stout!*
 
I can't wait til these guys set pods! Do you have any (High Performance Liquid Chromatography) HPLC equipment to do capsaicin analysis? If you did an assay across all your plants with an N=3 or N=5, you could get dope results on the efficacy of colchcine on increasing capsaicin in pods. Plant size, growth rate, pod production and size would be easy to quantify but I doubt you'd want to taste test the capsaicin content of these suckers. I'm really excited to see these results. 
 
Also, do you think measuring optical density of a capsaicin solution would work? You could definitely extract the capsaicin with methanol or hexane and then measure the OD and backtrack to calculate the concentration with the Beer-Lambert Law. If you don't have HPLC and you or your wife got an A in orgo, it wouldn't be too difficult. Just time consuming.
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I`m not convinced they will develop, to be honest. The root system development is severely compromised in all of them that look this way. Of all the seedlings I have, they either look normal, or they look like this one. I`ll probably have to go back and re-do the experiment at the same concentrations, but different treatment times.
 
I do not have access to HPLC anymore, no, although I think I spent half my life running bloody HPLC and it would be extremely simple to do capsaicin analysis. If you want to buy me one I`m sure ebay would have one for about $5-6k  :rofl:
 
I suspect that optical density measurements would only work if you had exceptionally pure solutions. Extracts from peppers would probably be far too complex and variable. I doubt UV or IR spectroscopy would be much use without some pretty fancy separation methods to purify the samples first. Even HPLC would probably require some sort of specialist extraction/fractionation techniques before injecting samples on to the column. 
 
You think another dose at around 8 inches tall or so would have been a good idea to ensure there are no more diploid cells and that it would have enough root to be okay?
 
sciurus, he made the mistake of using brown moruga for this one. He's darn well going to be taste testing these things.
 
I'd personally take equal weights/volumes of pepper extract and taste test small amounts of them to find out the difference, it would be the easiest.
 
just thinking out loud here. If natural tetraploid Capsicums are out there, wouldn't there also have to be some sterile (possibly seedless) triploids as the progeny of backcrosses with diploid stock? I haven't heard of anyone talking about their seedless peppers.
 
cruzzfish said:
You think another dose at around 8 inches tall or so would have been a good idea to ensure there are no more diploid cells and that it would have enough root to be okay?
 
sciurus, he made the mistake of using brown moruga for this one. He's darn well going to be taste testing these things.
 
I'd personally take equal weights/volumes of pepper extract and taste test small amounts of them to find out the difference, it would be the easiest.
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Don't get me wrong, I would pay cash to see a video of an N=3 pepper taste test assay with TBMs but I don't think Nigel would survive it!
 
I also don't think the human tongue is a a good way to test these. Even if you had a real knack for tasting peppers, your heat tolerance goes up in one sitting and spacing them out would force the tester to rely on memory which is unreliable. I doubt that the difference would be Jalapeno vs. TBM. I'd guess that if there is a heat increase, it's probably only by a few hundred SHU.
 

Ají hombre said:
just thinking out loud here. If natural tetraploid Capsicums are out there, wouldn't there also have to be some sterile (possibly seedless) triploids as the progeny of backcrosses with diploid stock? I haven't heard of anyone talking about their seedless peppers.
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Maybe triploidy simply doesn't result in sterility in Capsicum. Honestly, I have no idea since I haven't taken genetics yet. That said, maybe an effect of triploidy in Capsicum is part of a mechanism the plant needs to live. If that's true, maybe triploids don't even germinate or perhaps never set pods? The only way to tell is to wait for Nigel's nubby plants to bear fruit.
 
cruzzfish said:
Triploid shouldn't kill anything if everything is tripled. If it's just one, well then that plant is f*cked.
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I wouldn't think so, Maybe triploidy doesn't cause sterility in Capsicum? I would guess it produces one haploid and one diploid sex cell per miotic division so one in two pollinations would fail?
 
cruzzfish said:
You think another dose at around 8 inches tall or so would have been a good idea to ensure there are no more diploid cells and that it would have enough root to be okay?
 
sciurus, he made the mistake of using brown moruga for this one. He's darn well going to be taste testing these things.
 
I'd personally take equal weights/volumes of pepper extract and taste test small amounts of them to find out the difference, it would be the easiest.
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Maybe treatment of plants with a larger root system would be the way to go. This is the first pilot experiment, so I just chose the simplest method that had worked in many other situations. 
 
Not Brown Moruga, just regular Red Moruga (TSMB). I wanted to choose something very hot, but something that had nobodies name (or ego) attached to it. If you think about superhot peppers, there really aren`t that many choices. I also had a couple of hundred TSMB seeds. The mean for TSMB is around 1.2 million SHU, high is just over 2 million SHU. In all honesty, I`d rather see big bushy plants with apple-sized pods that any increase in SHU measurements. Make no mistake, though, any pods will be taste tested on camera!!!
Ají hombre said:
just thinking out loud here. If natural tetraploid Capsicums are out there, wouldn't there also have to be some sterile (possibly seedless) triploids as the progeny of backcrosses with diploid stock? I haven't heard of anyone talking about their seedless peppers.
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Seedless peppers do occur, but I expect not because they are triploid, as it`s often just a few on one plant, others having seeds. As you suggest, if there were tetraploids you might expect triploids in the cross with diploids. If that happened, as a plant breeder, you'd just throw away the plant, as it`s no use. The assumption that there are (or have been at some point) tetraploid Capsicums around is a fair one, but given the relatively short time breeding projects have been around, who knows if they exist or not in our populations. It does seem very possible, though. To the best of my knowledge, all Capsicums examined have either 24 or 26 chromosomes, with all cultivated types having 24. That does not mean polyploids don`t show up, but would anyone recognize one if it did? 1 plant in a field of 20,000? I very much doubt it.
 
A good example of a polyploid plant that has been selected for more than 10,000 years is wheat. There is diploid wheat, but the main wheat grown, "bread" wheat is actually sexaploid. Durum wheat, used to make pasta, is tetraploid. 
 
SciurusDoomus said:
I wouldn't think so, Maybe triploidy doesn't cause sterility in Capsicum? I would guess it produces one haploid and one diploid sex cell per miotic division so one in two pollinations would fail?
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Yeah. Half fertility rates. Odd chromosome numbers in general cause issues, but even tends to be a lot more reliable. I'm beginning treatment on already grown plants now.
 
cruzzfish said:
Yeah. Half fertility rates. Odd chromosome numbers in general cause issues, but even tends to be a lot more reliable. I'm beginning treatment on already grown plants now.
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Foliar feed? What was your colchicine source/concentration?
 
And Nigel, have you considered grafting a tetraploid stalk on a rugged diploid root stock? Would it have any chance of taking?
 
Once I get everything set up, I'm dunking my DWC plants in .05% for 24 hours and see what happens.
 
Grafting is a good idea, but these plants are small and therefore too hard to work with, at least that's what I'd think.
 
     I was thinking someone could grow a plant for a couple of weeks to get a good root system established, then chop it back to a couple of nodes. Apply colchicine solution to one node before a bud forms. That way you'd be able to visually compare the growth between treated vs. untreated nodes on the same plant and get an idea of whether anything happened.
     It seems (to me at least) like the colchicine would have a better chance of soaking into the tender young cells of a meristem rather than hoping it soaks into an entire plant via spraying an entire plant. Also, no grafting required.
 
Nigel have you tried doctoring the base of the plant with some rooting hormone to see if it would induce root growth? you could also give them a shot of Gibberellic Acid to give them a jumpstart on their growth.
 
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