THE MOST COMMON QUESTION
Question: The federal THC limit is 0.3% for industrial hemp. I’ve seen other seed companies claiming consistent results below that level. Will your seed produce a field of 0.3% THC plants?
The Mandate
The 0.3% THC mandate is a problematic interpretation and application of early cannabis science. In 1976, Canadian horticulturalists Ernest Small and Arthur Cronquist published an article in Taxon, entitled “A Practical and Natural Taxonomy in Cannabis,” in which they provide the seminal demarcation between “industrial hemp” vs “marijuana” that has been adopted by governments around the world. In their words:
The Detailed Answer
Figure 1. Whole Sequence Cannabis Genome Browser
Figure 2. Cannabis Chemotype Typology
Even with the THC genes “off,” type III and IV plants still produce a small amount of THC; the reason for this was unknown until we discovered the answer in early 2019. After tens of thousands of chemical and genetic tests (including whole genome sequencing of hundreds of plants), we now know that THC accumulation in type III and type IV varieties is due to CBCA genes, located on chromosome nine (remember: CBD and THC genes are located on chromosome six).
Figure 3. Cannabis Chromosomes
The CBCA synthase primarily produces trace amounts of CBCa, but, because of its 99% DNA sequence identity to the THCA synthase, it also produces similarly low levels of THCa. The latest research on these genes indicates that small variations in CBCA alleles (specifically, single nucleotide polymorphisms or SNPs) affect their conversion efficiency of the cannabinoid precursor CBGa to either CBCa or THCa. Additionally, the total copy number of CBCA gene loci play a critical role in determining final THC concentrations as well; after sequencing hundreds of type III plants, we have found that all have between 6-8 copies of the active allele and this number is directly related to ratio inheritance (and therefore total THC potential). This is not unique to our plant collection; rather, it is the case for all commercially available type III varieties. The ramifications of this finding should become the death-knell of the scientifically invalid 0.3% THC limit.
In our high cannabinoid hemp research programs, we have encountered plants that range from .05%-1% THC by dry weight on trimmed flowers. This means the federal limit of 0.3% is socially constructed rather than scientifically accurate (unless only leaves are tested!), but we must currently abide by this mandate, which makes sampling protocols and test-timing critical. High cannabinoid hemp farming is still possible (as evidenced by the explosion of this industry), but seed selection is also paramount for success. Farmers must consider a seed variety’s CBD:THC and/or CBG:THC ratio, total cannabinoid content, and appropriate sample timing in their state to achieve good results, as all three of these variables play critical roles in successfully growing a full-term crop of legal, industrial hemp.
Critical Considerations for Success
Ratio
For type III plants, the competitive rate of conversion between active CBDa and active CBCa genes is expressed as the proportion of CBD to THC present in the total cannabinoid content and is commonly referred to as “the ratio” (i.e. ratio of CBD to THC, 20:1, 35:1, 40:1, etc). The ratio of CBD:THC is fixed in each plant once it matures out of its juvenile phase and can be accurately detected by chemical analysis of new leaves after a month of growth (4th node minimum). Final, total THC is a function of total cannabinoid content and ratio:
As described above, there are no type III CBD varieties in existence today that breed true for ratios over +/- 30:1. Many seed vendors claiming otherwise will provide either (1) valid test results of one plant with a higher than 30:1 ratio and claim that the trait is stable or (2) fraudulent test results. True-breeding, ultra-high ratio varieties (50:1+), so far, have only been validated in our pure type IV CBG varieties.
Total Cannabinoid Content
“Cannabinoid content” refers to the final, maximum amount of total cannabinoids present in a plant (or population of plants if using an average). This is one of the single most important economic qualities of cannabis lines, one of the most widely manipulated results by unscrupulous breeders, and a key determinant for meeting federal THC compliance levels. High cannabinoid content seed lines separate the “winners” from the “losers” in hemp production; if oil production is your goal, the low cannabinoid, $20 per pound seed will get you nowhere. Finola is a good example of this; many farmers love the low price of this seed, but plants average 1%-2% CBD under the best circumstances, making it cost prohibitive as a supply stock for extractors—tons of this flower material is composted each year due to its low marketability. Knowing the importance of high cannabinoid lines, some breeders claim outrageously high figures for their seed and offer no analytical evidence to back it up; others submit trimmed flower test results from their most stellar progeny and claim it’s representative of the entire population. While we would love to have a field full of 25% CBD plants grown from seed (as some advertise for $14 each!), we know that this is logically impossible—caveat emptor. We strive to provide accurate information on our varieties rather than marketing hype and to do so in a transparent, reproducible fashion. Compared to clones, farmers growing industrial hemp from seed will be confronted with more variation in total cannabinoid content, but predictably so. Cannabinoid content is a polygenic, but heritable trait; it is derived from the complex interaction of many different genes and protein expression networks, but is consistently passed on with equal contributions from both parents at the population level. The average cannabinoid content in a population of F1 sibling plants is (generally) the average of the parents’ total cannabinoid production (true heterosis—i.e. hybrid vigor—is an exception to this). Consider the following hypothetical pairing of parent plants P1 (20% cannabinoid content) and P2 (10% cannabinoid content):
Figure 4. Cannabinoid Total Inheritance in F1 from Sources P1 and P2
The F1 progeny (i.e. offspring) of parents P1 and P2 will average 15% total cannabinoid content. While this allows for some predictability on financial returns (since total cannabinoid content is what sets the price for your product as a farmer), it does not mean that each individual plant will be 15%. Total cannabinoid content in seed lines are “normally distributed,” which means that most plants (68%) will be within one standard deviation of the population mean, some plants (27%) will be within two standard deviations of the population mean, and a few (5%) will be three or more standard deviations away from the mean. Returning to our hypothetical example above and using a standard deviation of 1%, a field of F1 plants derived from our P1 and P2 parents would follow the distribution presented in Figure 5. Distribution of Cannabinoid Totals in F1 Population
Sampling Protocol and Timing--Whose 0.3% THC Counts?
With good quality, high cannabinoid type III CBD seed, compliance is determined by the timing of test, sampling protocol, and laboratory accuracy. For the past five years of hemp production (authorized by the 2014 Farm Bill), every state has implemented their own sampling and testing procedures; this has led to a confusing patchwork of policies where plants certified as "hemp" in one state may not pass compliance testing in another state. Examining Oregon, Kentucky, and Pennsylvania rules provides an illuminating example.
Timing of Test
- Oregon: 28 days before harvest begins
- Kentucky: 15 days before harvest is complete
- Pennsylvania: 70-80 days after planting
Cannabinoid content increases once flower formation commences; the most significant increases occur between weeks 4 and 6, then slowly accumulate until peak ripeness (i.e. harvest) around 56-60 days into flowering. An identical plant grown in separate locations (Oregon, Kentucky, and Pennsylvania--see Figure 6) would be tested at very different times in their developmental process; this obviously poses a challenge to plant breeders, farmers, law enforcement, and state regulators alike--particularly when considered in relation to the underlying scientific invalidity of the 0.3% THC rule.
Figure 6. Cannabinoid Content Over Eight Weeks of Flowering With State Compliance Test Timing
Sampling Protocol
- Oregon: Top 8 inches of 30 plants; delta-9 only (no decarboxylation); carried out by private, nationally accredited labs
- Kentucky: Top 20cm of 5 plants; delta-9 THC + (THCa * 0.877); carried out by Kentucky Department of Agriculture
- Pennsylvania: Terminal flowers only (top 2 inches); delta-9 THC post-decarboxylation; farmer collects sample, sends to any lab in the US
Inter-lab Variability
We held a lab "test-off" at our R&D field in 2018 on a day neutral CBD variety we developed ("RNA"); every nationally-accredited (NELAP) cannabis lab in Oregon was invited to participate. All labs were required to follow the state mandated sampling methodology and testing protocol, and cut samples simultaneously on the same day--28 days before harvest. The results are summarized in Figure 6 below.
Figure 6. Pre-Harvest Compliance Test Results from Operational 3rd Party Labs
Samples were collected from 55,750 plants over 12 acres of undulating field conditions by 8 separate sampling teams following ODA’s prescribed pattern and protocol. Significant differences in price, LOQ levels (lower is better, when accurate), and gross total content were apparent. All of our samples came in well below the total 0.3% THC threshold, with the exception of Cascadia’s test (which was grossly mishandled by commingling our sample with a high THC sample after accidentally throwing both in the trash (you can't make this stuff up)--the company no longer operates in Oregon). Sampling technique (i.e. biomass selection) has the greatest influence on total cannabinoid content (both CBD and THC) and this is apparent when comparing results; for reference, the reported THC content from Green Leaf Labs, Pixis, and SC Labs are the closest to those we have received from ODA in previous compliance events with this particular variety. Not all labs test for the acidic forms of non-THC and CBD cannabinoids; at least one lab (Evio) inappropriately reported high levels of CBN instead of CBGA, while missing CBCA accumulations entirely and misreporting their actual LOQs. While this does not affect your THC compliance, it is definitely an issue that savvy growers should pay attention to, since many states place civil responsibility on farmers to select a lab with great record-keeping practices (Oregon requires growers to maintain records for at least 3 years post-harvest—always keep copies for yourself, just in case).
Figure 6 also includes imputed CBD:THC ratios derived from the test-day results. As we have previously described, these ratios represent important breeding markers for those interested in maximizing oil production, since higher ratios allow for greater CBDa in ultra-low THCa, federally compliant plants. In these tests, results ranged from 16:1 to over 56:1 amongst labs, with an average (mean) of 27.2:1. We are not surprised to find that the lab with the lowest LOQs (Pixis) produced results in agreement with this population average. This is an important finding for farmers to consider when vetting claims made by seed vendors or breeders—there is no such thing as “non-detectable THC” in CBD-rich hemp just yet, only inaccurate lab results.
