Cubing

GrapeStreet · Feb 25, 2011

A collection of article briefs regarding the breeding technique, "Cubing."

Cubing works on the basis of mathamatical probabilities with respect to gene frequencies. The more males you use with each cross, the better the chance that your reality matches the theory. In theory, with the first backcross, 75% of your genepool will match the genepool of the P1 parent being cubed. Squaring increases this to 87.5% and cubing increases it to 93.75%. You can arrive at these numbers by taking the average between the two parents making up the cross. For instance, you start by crossing the P1 mom (100%) with and unrelated male (0%) getting 100% + 0% divided by 2 = 50%. Therefore, the offspring of this first cross are loosly thought of as being 50% like the mom. Take these and do your first backcross and you get 100% (mom) + 50% divided by 2 = 75%. And this is where we get the 75% for the first backcross. Same thing applies as you do more backcrosses. As you will see later, you can apply this same probability math to specific genes or traits, and this can have a dramatic effect on your methodology and selection methods.

Your selection of the right males for each backcross are the crucial points for success with this technique. In each case, you could select males that contain the genes you want, or you could inadvertedly pick those individuals that carry the unwanted recessive genes. Or more likely, you could just pick individuals that are heterozygous for both genes like the P1 mom being backcrossed. The easiest way to deal with this is to start by only looking at one gene and one trait, like lets assume that flavour is determined by a single gene (in reality it's probably not). And do some punnet squares to show gene frequencies through 3 generations of backcrossing. Now lets assume that we found a special pineapple flavoured individual in our pine flavoured population that we wanted to keep. The gene causing the pineapple flavour could be dominant or recessive and the selection abilities and cubing outcome will be different in both cases.

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In reality, most of the traits we select for like potency are influenced by several traits. Then the math gets more complicated if you want to figure out the success rate of a cubing project. Generally speaking, you multiply the probabilities of achieving each trait against each other. For example, if your pineapple trait was influenced by 2 seperate recessive genes, then you would multiply 87.5% * 87.5% (.875 * .875 *100) and get 76.6%. This means that 76.6% of the offspring would be pineapple flavoured. Now lets say the pineapple trait is influenced by 2 recessive traits and and a heterozygous dominant one. We would multiply 87.5% by 87.5% by 71.9% (.875*.875*.719*100) and get 55%. Just by increasing to three genes, we have decreased the number of cubed offspring having pineapple flavouring down to 55%. Therefore, cubing is a good technique where you want to increase the frequency of a few genes (this is an important point to remember ), but as the project increases, the chance of success decreases .... at least without some level of selective pressure.
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CCubing in reality is less controlled than this. Cubing is a way of increasing the frequency in a population for a certain trait. It MAY not result in true breeding but it will promote a trait in a bunch of plants. Also the actual selection process is somewhat random.
In a population we select a mother plant that we want to keepbecause of her features. In the same population we gather pollen from50% of the males that have characteristics similar to the mother plant and 50% that do not. The pollen is mixed in their respective portions. So we have two packets of pollen in the end. We must clone the female to create 2 females. We then use the 2 packets of pollen on each clone separately.

When we grow out the offspring from the two females we will select the population of the offspring that mostly resembles the mother plant traits that we are looking to promote in the population. What happens is that the best male pollen should have been selected by the female as the one that she prefers. The reason for taking the 2 sets of pollen from 2 sets of males is to create a control experiment to show how this method actually interferes with the frequencies of the gene pool. By right, your selection of poor male pollen will only bring about
a poorer quality population that do not resemble the female clone. In reality we only select pollen from the best males that most resembles the female when we use this method.

Do you remember one of the laws that breaks equilibrium?Non-random mating and Natural selection.Well that is what we are doing here. The resulting offspring should have a high frequency for the traits that we like in the mother plant. The problem is that we do not know which male from the bunch is the one that WON this female over, but it is quicker and less time consuming in creating a strain that is somewhat uniform for a specific trait. By repeating the process we can help increase the mother plants trait frequency in the offspring's population but we will probably end up with some plants that are non-uniform for that trait. The best way to achieve this process is as follows. Find a female that you like and clone this female and her male parent. Take pollen from the male parent and pollinate the female. The offspring should contain a 50%/50% of the genes for both parents. Take pollen from the males of that offspring and mix them together. Pollinate a clone of the mother. This step should insure that selection is no longer random and you are promoting the frequency of the mother's traits in the next offspring. Repeat the process two more times and you will have effectively CUBED (meaning backcross x 3) this strain. This can push the mother plants traits as high as 90% in a population but we will probably get some non-uniform plants in the offspring too. Cubing does not really help us to select for traits that we want, like in our Silver Kush experiment. It simply helps us to keep a few traits that a mother plant has. Cubing is a common procedure adopted by breeders who find a good healthy mother plant in a selection of seeds that someone has given them. This method can also fail very quickly if your selection of males are the wrong choice.
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"you've just discovered the biggest myth (IMNSHO) of marijuana breeding- it is a mistake that almost EVERYONE makes (including many of the most respected breeders!).

Backcrossing will not stabilize a strain at all- it is a technique that SHOULD be used to reinforce or stabilize a particular trait, but not all of them.

For e.g.- G13 is a clone, which I would bet my life on is not true breeding for every, or even most traits- this means that it is heterozygous for these traits- it has two alleles (different versions of a gene). No matter how many times you backcross to it, it will always donate either of the two alleles to the offspring. This problem can be compounded by the fact that the original male used in the cross (in this case hashplant) may have donated a third allele to the pool- kinda makes things even more difficult!

So what does backcrossing do?
It creates a population that has a great deal of the same genes as the mother clone. From this population, if enough plants are grown, individuals can be chosen that have all the same traits as the mother, for use in creating offspring that are similar (the same maybe) as the original clone.
Another problem that can arise is this- there are three possibilities for the expression of a monogenic (controlled by one gene pair) trait.

We have dominant, recessive, and co-dominant conditions.

In the dominant condition, genotypically AA or Aa, the plants of these genotypes will look the same (will have the same phenotype, for that trait).

Recessive- aa will have a phenotype

Co-dominant- Aa- these plants will look different from the AA and the aa.

A perfect example of this is the AB blood types in humans:

Type A blood is either AA or AO
Type B blood is either BB or BO
Type AB blood is ONLY AB
Type O blood is OO.

In this case there are three alleles (notated A, B, and O respectively).

If the clone has a trait controlled by a co-dominant relationship- i.e. the clone is Aa (AB in the blood example) we will never have ALL plants showing the trait- here is why:

Suppose the clone mother is Aa- the simplest possibility is that the dad used contributes one of his alleles,
let us say A. That mean the boy being use for the first backcross is either AA or Aa. We therefore have two possibilities:

1) If he is AA- we have AA X Aa- 50% of the offspring are AA, 50% are Aa. (you can do the punnett square to prove this to yourself).

In this case only 50% of the offspring show the desired phenotype (Aa genotype)!

2) If the boy being used is Aa- we have Aa X Aa (again do the punnett square) this gives a typical F2 type segregation- 25% AA, 50% Aa, and 25% aa.
This shows that a co-dominant trait can ONLY have 50% of the offspring showing the desired trait (Aa genotype) in a backcross.

If the phenotype is controlled by a dominant condition- see example #1- all 100% show the desired phenotype, but only 50% will breed true for it.

If the phenotype is controlled by a recessive condition- see example #2- only 25% will show the desired phenotype, however if used for breeding these will all breed true if mated to another aa individual.

Now- if the original dad (hashplant) donates an 'a' allele, we only have the possibilities that the offspring, from which the backcross boy will be chosen, will be either Aa or aa.
For the Aa boy, see #2.
For the aa boy (an example of a test cross, aa X Aa) we will have:
50% aa offspring (desired phenotype), and 50% Aa offspring.

Do you see what is happening here? Using this method of crossing to an Aa clone mother, we can NEVER have ALL the offspring showing the desired phenotype! Never! Never ever ever! Never!! LOL

The ONLY WAY to have all the offspring show a Aa phenotype is to cross an AA individual with an aa individual- all of the offspring from this union will be the desired phenotype, with an Aa genotype.

Now, all of that was for a Aa genotype for the desired phenotype. It isn't this complicated if the trait is AA or aa. I hope this causes every one to re-evaluate the importance of multiple backcrosses- it just doesn't work to stabilize the trait!

Also- that was all for a monogenic trait! What if the trait is controlled by a polygenic interaction or an epistatic interaction- it gets EVEN MORE complicated? AARRGH!!!!

Really, there is no need to do more than 1 backcross. From this one single backcross, as long as we know what we are doing, and grow out enough plants to find the right genotypes, we can succeed at the goal of eventually stabilizing most, if not all of the desired traits.

The confusion arises because we don't think about the underlying biological causes of these situations- to really understand this; we all need to understand meiosis.

We think of math-e.g. 50% G13, 50% hashplant

Next generation 50% G13 x 50% g13hp or (25% G13, 25%HP)

We interpret this as an additive property:
50% G13 + 25% G13 +25% HP = 75% G13 and 25% hashplant

This is unfortunately completely false- the same theory will apply for the so called 87.%% G13 12.5% HP next generation, and the following 93.25% G13, 6.25% HP generation; we'd like it to be true as it would make stabilizing traits fairly simple, but it JUST DOESN'T work that way. The above is based on a mathematical model, which seems to make sense- but it doesn't- we ignore the biological foundation that is really at play.

I hope this was clear, I know it can get confusing, and I may not have explained it well enough- sorry if that is the case, I'll try to clear up any questions or mistakes I may have made.

Have fun everyone while making your truebreeding varieties, but just remember that cubing (successive backcrosses) is not the way to do it!

-Chimera"
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Wow that is painful for me to read.... I must have been pretty stoned when I wrote that, now years ago. I'm glad at least that it's info that has spread a little knowledge and perspective into the community over the years.. I do however need to re-write it...

Here's a snipet on backcrossing from the breeding chapter I wrote for Jorge's most recent version of the bible.

Backcross Breeding â€“
A type of breeding that involves repeated crossing of progeny with one of the original parental genotypes; cannabis breeders most often cross progeny to the mother plant. This parent is known as the recurrent parent. The non-recurrent parent is called the donor parent. More widely, any time a generation is crossed to a previous generation, it is a form of backcross breeding. Backcross breeding has become one of the staple methods clandestine cannabis breeders use, mainly because it is a simple, rapid method when using greenhouses or grow
rooms, and requires only small populations. The principle goal of backcross breeding is to create a population of individuals derived mainly from the genetics of one single parent (the recurrent parent).

The donor parent is chosen based on a trait of interest that the recurrent parent lacks; the idea is to introgress this trait into the backcross population, such that the new population is comprised mainly of genetics from the recurrent parent, but also contains the genes responsible for the trait of interest from the donor parent.

The backcross method is a suitable scheme for adding new desirable traits to a mostly ideal, relatively true-breeding genotype. When embarking on a backcross breeding plan, the recurrent parent should be a highly acceptable or nearly ideal genotype (for example, an existing commercial cultivar or inbred line). The ideal traits considered for introgression into the new seed line should be simply inherited and easily scored for phenotype. The best donor parent must possess the desired trait, but should not be seriously deficient in other traits. Backcross line production is repeatable, if the same parents are used.

Backcross breeding is best used when adding simply inherited dominant traits that can easily be identified in the progeny of each generation (example 1). Recessive traits are more difficult to select for in backcross breeding, since their expression is masked by dominance in each backcross to the recurrent parent. An additional round of open pollination or sib-mating is needed after each backcross generation, to expose homozygous-recessive plants. Individuals showing the recessive condition are selected from F2 segregating generations and backcrossed to the recurrent parent (see example 2).

Example 1â€“ Backcrossing: Incorporating a dominant trait

Step1â€“ Recurrent Parent x Donor Parent
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V
F1 Hybrid generation

Step 2 â€“ Select desirable plants showing dominant trait, and hybridize selected plants to recurrent parent. The generation produced is denoted BC1 (some cannabis breeders break from botanical convention and denote this generation Bx1. BC1= Bx1).

Step 3 â€“ Select plants from BC1 and hybridize with the recurrent parent; the resulting generation is denoted BC2.

Step 4 â€“ Select plants from BC2 and hybridize with the recurrent parent; the resulting generation is denoted BC3.
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Example 2 Backcrossing: Incorporating a recessive trait

Step1â€“ Recurrent Parent x Donor Parent
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V
F1 Hybrid generation

Step 2 â€“ Select desirable plants, and create an F2 population via full sib-mating.

Step 3 â€“ Select plants showing the desired recessive trait in the F2 generation, then hybridize selected F2-recessive plants to the recurrent parent. The generation produced is denoted BC1.

Step 3 â€“ Select plants from BC1, and create a generation of F2 plants via sib-mating; the resulting generation can be denoted BC1F2

Step 4 â€“ Select desirable BC1F2 plants showing the recessive condition, and hybridize with the recurrent parent; the resulting generation is denoted BC2.

Step 5 â€“ Select plants from BC2, and create an F2 population via sib-mating; denote the resulting generation BC2F2.

Step 6 â€“ Select plants showing the recessive condition from the BC2F2 generation, and hybridize to the recurrent parent; the resulting generation is denoted BC3.

Step 7 â€“ Grow out BC3, select and sib-mate the most ideal candidates to create an F2 population, where plants showing the recessive condition are then selected and used as a basis for a new inbred, or open-pollinated seed line.

This new generation created from the F2 is a population that consists of, on average, ~93.7% of genes from the recurrent parent, and only ~6.3% of genes leftover from the donor parent. Most importantly, one should note that since only homozygous-recessives were chosen for mating in the BC3F2 generation, the entire resulting BC3F3 generation is homozygous for the recessive trait, and breeds true for this recessive trait. Our new population meets our breeding objective. It is a population derived mainly from the genetics of the recurrent parent, yet breeds true for our introgressed recessive trait.

Backcross derived lines are expected to be well-adapted to the environment in which they will be grown, which is another reason backcrossing is often used by cannabis breeders who operate indoors. Indoor grow rooms are easily replicated all over the world, so the grower is able to grow the plants in a similar environment in which they were bred. Progeny therefore need less extensive field-testing by the breeder across a wide range of environments.

If two or more characters are to be introgressed into a new seed line, these would usually be tracked in separate backcross programs, and the individual products would be combined in a final set of crosses after the new populations have been created by backcrossing.

The backcross scheme has specific drawbacks, however. When the recurrent parent is not very true-breeding, the resulting backcross generations segregate, and many of the traits deemed desirable to the line fail to be reproduced reliably. Another limitation of the backcross is that the â€œimprovedâ€ variety differs only slightly from the recurrent parent (e.g., one trait). If multiple traits are to be introgressed into the new population, other techniques such as inbreeding or recurrent selection may be more rewarding.

Hope that's a little more clear......
Respectfully,
-Chimera
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Briar Patch · Mar 3, 2011

Can't rep you anymore. Nice read, thanks for sharing.

Rippa · Mar 22, 2011

really good information....

Loner Stoner · Apr 25, 2011

Ok, so I was doing some Pummett squares to see if backcrossing was truly neccesarry. I'll only be using two alleles, but please try to follow my steps to tell me if I'm thinking of this correctly.

Desired Trait: Denser buds

Female plant
First allele: A (we'll say it's a gene for denser buds)
Second allele: a (let's say it's a gene for less dense buds) <--- probably bad examples

Male plant
First allele: A
Second allele: a

So after doing some Pummett squares I came up with four possible genotypes, with two possible phenotypes.

Genotype possibilities:
AA, Aa, Aa, aa (25% Dominant, 50% Co-Dominant, and 25% recessive)

Phenotype possibilities:
AA, Aa - Denser buds (Should be around 75% of population of offspring)
aa - Lanky Buds (should be around 25% of population of offspring)

Ok, this is where I started getting confused. I didn't really understand the importance of backcrossing, so when I went through this process in my head I didn't think to clone the original mother (since I was trying to isolate a single phenotype, instead of trying to copy all the phenotypes of the mother itself, or a more uniform offspring) So anyway, for the next step, I assigned a male of female designation to each genotype randomly (given male to female ratio in cannabis is 1:1)

Since the aa genotype did not have the desired trait of denser buds, I removed all of those types from the breeding project. So I was left with roughly 66% Aa genotype, and 33% AA genotype based on the remaining offspring from the original mother and father breeding.

This is where my method got extremely difficult, because it would require a lot of growings. I would randomly breed the pollen from a single male to a single female (instead of mixing all the pollen up with different males). Theoretically if I got lucky, a AA male will breed with an AA female. All offspring would be AA, therefore having the desired trait. However, since there are more Aa present in the original offspring, there are two other possible combinations I could create, being AA x Aa and Aa x Aa.

With AA x Aa I'll have more possible AA, since due to some more Pummett square I determined we'd have about 50% AA and 50% Aa. This increases the chance of me matching up a AA with an AA using the AA x Aa offspring versus the Aa x Aa offspring.

With Aa x Aa I'd have only a roughly 25% chance of producing an AA genotype. This will produce the same exact results (roughly) produced by combining the original parents.

So as you can see, I can achieve the AA x AA the second breeding so long as there is no aa genotype. It has to be at least co-dominant. The reason I did a second breeding amongst the first offspring was to increase the chances of an AA genotype. I could have backcrossed I suppose, but if the parent plant is an Aa, I'd be allowing that recessive a to be passed on to at least some of the population depending on the male genotype.

Now comes the most crucial part of this process being successful. You have to seperate each different set of breedings. Once you have your first set of seeds from the Aa female and Aa male which we'll call G1 (generation 1 - the seeds), and you've removed the obvious aa genotypes, and you've bred random males with random females (not just a big mix of pollen. Just like humans do it... one man per woman, one woman per man) When all your new G2 babies grow up and reveal their phenotypes, do the same steps again breeding random males with females. What we want are the AA x AA matchups. The only way we know if we succeded or not is to backcross to the original mother (ok damn I just realized why backcrossing is important... which disproves my whole theory. But I've been on this for hours and I can't just delete this post after all the time i spent on it... Damn I'm stupid stoned.

Was I at least on the right track?? Lol. My head hurts now. Off to burn one. Ugh but I need to understand how this works... anyone think they can put my mind back on the right track? Cuz I really thought I had something there...

Basically I want the AA x AA. That I know for sure! Tricky thing is knowing if it's an AA x AA, or something like a Aa x AA..

Hmm... Super stoner comeback!

What if for every male I bred with a female, I seperated it from the others. So with the original Aa x Aa, and having 25% AA, 50%Aa, 25%aa, I'd remove the aa, what I would do is breed randomly, and grow out each child until I had a seedstock that was all possessing the desired trait. So let's say I happened to match up an AA male with an AA female from G1. I would be able to confirm that in G3 by growing out the G2 seeds, which would ALL have the desired trait, right? Then to further varify to make sure I don't have an AA x Aa hybrid, I grow out a G3(Third generation). IF was truly an AA x AA I'd have succeeded. If the male-female coupling wasn't an AA x AA match, then I'll have some recessive dominant genes showing (such as the less dense lanky buds), or all the recessive genes could just be hiding, but will reveal themselves in the third generation of inbreeding.

Whew, I think I made sense. I hope so guys, cuz I was trying to do this having only taken a basic genetics course at my university. And this is all just for one trait (dense buds). We have our work cut out for us if we want to isolate multiple traits, or are dealing with a third allele as the OP mentioned.

So anyway, the whole point of this semi-educational (I say that because I don't know if I'm correct or not) digression was to prove that backcrossing was unneccessary in isolating desired traits.

A quick review of the whole process:

Male is Aa , A being the desired trait
Female is Aa

Document every breed!

Aa x Aa produces AA, Aa, aa. This is G1 (1st Generation)
Remove the aa.

Match up random G1 males with random G1 females. Produces G2 seeds
Grow the G2 seeds
Seperate every G2 specimin.

G2 specimens will either be AA x AA (we want) or AA x Aa (dont want) or Aa x Aa (worst)

Lol, again I find the importance in backcrossing. To verify if your G2 specimen is indeed an AA x AA, you'd have to backcross the G2 specimen with the G1 parent of opposite sex. Produces G3 seeds
If you successfully matched up an AA with an AA in G1, then G3 specimins will ALL have the dominant traits. However if was an AA with an Aa, then there will be undesireable traits in G3.

If unsuccessful then you trash that plant. ITs useless. If it was successful then obviously you keep it, and clone it, or set it to seed. You're more likely to have an AA x AA if you create more specimens back in G1.

This process is difficult because it takes three generations to confirm if you have an AA x AA. But like I said, you'll have tons of specimens going at once, and you just keep the ones that you know are the AA x AA through G3 verification.

So experts, was my process correct? I guess I didn't disprove the need of backcrossing, but I didn't do it until the very end. I guess same result different process?

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