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Old 03-12-2018, 10:54 AM #411
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Originally Posted by socioecologist View Post
Always best to self to reduce heterozygosity. 25% will be high CBD (type III), 50% will be 1:1 (type II), and 25% will be high THC (type I). Where:

T = active THC allele
t = inactive THC allele
D = active CBD allele
d = inactive CBD allele
Thanks for the info and chart. I appreciate it.

OK, so, it's best to reverse sex and self pollinate, and then 50% of the offspring will be the same as the parent, which should be 1:1.

Correct?

(When i mentioned that the CBD Crew seeds were 'high CBD' i actually meant 1:1 CDB:THC, were you aware that's what i meant?)

I just want offspring that is the same as the parent.

I think i read somewhere that we could easily test for CBD levels at home somehow, is this just a false memory i have?

Thanks again for any help.
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Old 03-12-2018, 07:24 PM #412
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...they are 8 centromeres apart on the same chromosome, but they readily segregate...
I re-read the publication with the genetic map: They published 1.1 cM which corresponds to a hypothetical 1.1% uncoupling/segregation rate although at <7 cM segregation is fairly unlikely. Stupid thing is that there are hotspots where crossovers occur and where not. The chance of segregation derived from breeding experiments is given as LOD value: If I'm reading the graph correctly, the LOD for THCA/CBDA is 40 which means that segregation occurs only in 1 of '1 followed by 40 zeroes' meiosis. Hence, chances to obtain a double homozygous plant either of the BT/BD or Bt/Bd type in the F2 is at 10^40 times 10^40 or simply said never. But all the genes on the chromosome LG6 have values greater than 3 which means that there basically never occurs any crossing over on the whole chromosome! Or I'm reading something wrong (I admit, I never really learned how to used/read gene maps and only did/do it for fun)???
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Old 03-13-2018, 02:43 PM #413
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I was referring to something else .
At the current state of scientific error, talking of TTdd and ttDD in case of drug type cannabis and hemp, respectively, is correct but using a 4x4 Punnett square isn't. The Td and tD alleles are linked and hence a simple 2x2 square with simplified TD x TD suffices.

THCA and CBDA are of (co-)dominant and not intermediate inheritance (one bell shaped distribution of cannabinoid quantity in the F2 generation, as shown in several publications) and therefore cannabinoid amount is rather due to other traits (available precursors, trichome density etc.). This means that chemotype testing (e.g. chromatography) won't tell genotype ergo distinguish between homo- and heterozygous plants: TTdd and Ttdd will both have THC and no CBD whereas TTDd, TtDd, TtDD, and TTDD all have THC and CBD and that probably at a similar ratio.
Furthermore, your Punnett square not only breaks down to each 3 plants with only THC or CBD and 9 with both cannabinoids (i.e. 1:3:1) but there's also 1 plant with neither THC nor CBD and hence high CBG. That's not what's commonly seen in cross-breeding drug type cannabis with hemp or simply breeding with intermediate types such as charas or hash varieties. Supposedly, there's never been even an indication in any of the germplasm analyses and cross-breeding experiments for the appearance of a double homozygous individual neither of the TTDD nor the ttdd type. Treating the BT and BD loci as one single B locus (i.e. 1:2:1) is closer to reality. Although, none of the publications I've read really hit it spot on. There's always some discrepancy especially the slight shift towards a higher CBD/THC ratio and a loss in overall cannabinoid content but that's the normal discrepancies between biology and statistics.

Taking your CBG variety as an example: chemotype testing won't tell you if it's a ttdd double homozygous plant, a loss of function mutation (either of the BT or BD allele), or the postulated B0 allele. Gene sequencing would be the only way to tell... Talking of which: the Phylos project might already hold the answer to that question.
Thanks OO for being a good sport, more-than-competent scientist, and gracefully putting up with a moment of acerbity from me. I appreciate your attention to detail and the effort you have exerted to dig into the truth here. This is a topic that I have been actively investigating for 3 years now from both academic and on-the-ground production perspectives. We are both wrong and both right, but the beauty of the exchange is that it is pushing forward our collective understanding of how reality is organized--that's the best part about "science"!

I was wrong above to say that the alleles "readily segregate". That's simply not true. However, they do, in fact, segregate and it is not 1 in 10,000,000,000,000,000,000,000 ,000,000,000,000,000,000 odds. You are correct that it is based on distance, but the rate of recombination is 1% per map unit (cM). It's not exceedingly rare for these events to occur when you are working with large populations. Take a look at page 1247 of the Weiblen et al. paper.; it happened in their n=62 sample.

Quote:
Genotyping also identified a single recombinant individual that was homozygous for the hemp-type THCA synthase and homozygous for the marijuana-type nonfunctional CBDA synthase.
I emailed George about this gem in Nov. 2016, as we were searching for pure CBG plants via ttdd (or more appropriately, td / td) and I inquired about the cannabinoid data on that individual plant. His data manager's response:

Quote:
Across 62 F2 individuals analyzed in the study, the CBG values ranged from 0.005 to 0.45; total cannabinoids ranged from 1.24 to 8.09. Respective values for CT132 do not seem to stand out in that regard. Among drug-type F2s in the study, log(%THC/%CBD) ranged from 1.09 to 3.13 putting CT132 towards the lower end of the range.

To clarify, CT132 is putatively recombinant in that it lacked detectable THCA1 synthase sequence whilst also being homozygous for non-functional CBDA synthase. However, it did exhibit full-length hemp-type THCA sequence bearing non-synonymous mutations with respect to THCA1 synthase. Lacking a detectable functional CBDA synthase, we wouldn't necessarily expect any expressed (truncated) CBDA synthase-like proteins in CT132 to functionally out compete an intact (full-length) THCA synthase even though the latter would be divergent in AA sequence from THCA1. Reiterating the statement in our paper, we interpret CT132's putative recombinant genotype to imply that in the absence of functional CBDA synthase competing for available CBGA, the AA-divergent but full-length hemp-type THCA synthase is a functional enzyme yielding the drug phenotype.
So no, it didn't lead to CBG--it was, technically, a type I phenotype with a type IV genotype. We run into similar plants in our CBG breeding program, though our alleles have been selected for their relative inefficiency at converting CBGa, so ratios of CBG to THC for these types range from 4:1 to 20:1. We are convinced that to get "pure" CBG (> 100:1) requires the presence of an additional gene. If I'm wrong about that, in my defense, it's data-driven (inductive) rather than theoretical (deductive). Oh, and phuck Phylos. .

The last bit I would like to offer is that, yes, in fact, ratios of CBD to THC (and vice versa) can be used as predictors of underlying genotype. It's not perfect due relative precursor conversion efficiency differences between allele variants, but it's damn obvious when combing through F2 progeny. For type III breeding we toss everything that is less than 20:1 CBD to THC...and do so for a very specific reason that is backed up with a lot of data.
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Old 03-13-2018, 07:40 PM #414
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Thanks for pointing out the page 1247 part, completely zapped it. First, I was all excited that it's 1% (which is quite a lot, considering) and then I got all disappointed when looking at the LOD...

VERY interesting quote too! New hope awakens.

Besides, I'm all yours: Science or experience alone have only a friction of the power they unfold when put together. We as a human race aren't strong because some individuals piss further, have deeper pockets, or a bigger whatsoever, no, it's because we can work and talk together, learn from one another, and contribute with our strength where others have weaknesses. Nobody's perfect but when we take the good parts of each one of us and put them together instead of bickering about mistakes, we as a community could well be. But not all are willing to go the extra mile, let alone for free...
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Old 05-14-2018, 05:37 PM #415
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Old 09-16-2018, 04:09 PM #416
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Originally Posted by Only Ornamental View Post
I re-read the publication with the genetic map: They published 1.1 cM which corresponds to a hypothetical 1.1% uncoupling/segregation rate although at <7 cM segregation is fairly unlikely. Stupid thing is that there are hotspots where crossovers occur and where not. The chance of segregation derived from breeding experiments is given as LOD value: If I'm reading the graph correctly, the LOD for THCA/CBDA is 40 which means that segregation occurs only in 1 of '1 followed by 40 zeroes' meiosis. Hence, chances to obtain a double homozygous plant either of the BT/BD or Bt/Bd type in the F2 is at 10^40 times 10^40 or simply said never. But all the genes on the chromosome LG6 have values greater than 3 which means that there basically never occurs any crossing over on the whole chromosome! Or I'm reading something wrong (I admit, I never really learned how to used/read gene maps and only did/do it for fun)???
I took this post as truth as it stands in cannabis for too long. No fault of OO, just a solid example of why asking "why?" and pushing forward to discover for ourselves is how progress unfolds. We have finished our summer synthase recombination experiments and the real rate is what you would expect to see based on their actual published distance. Not rare at all if you are used to large populations. In fact, frequent enough that it allowed us to be picky with which recombined mutants we kept for further breeding. With that out in the public now, I will also add a caveat: it's not so much about the recombination events as your starting alleles. Combining any of the known inactive alleles (i.e. td / td), doesn't produce a super useful chemotype. Magic can be made with novel alleles though.

Great example of how science works in real life right here, thanks for contributing OO!
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Old 09-17-2018, 10:12 AM #417
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Interesting discussion guys. Is that what they call "linkage disequilibrium"?

Do you ever do any functional research on how efficiently each of the many BT/BD alleles are able to catalyze the reaction? Affinity to substrate (CBG) and enzyme kinetics and what not.
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