Sire Lines & "Y" They Matter

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The THCAS and CBDAS share a high sequence similarity of 83% based on which a common ancestor was suggested initially (Taura et al., 2007b). Upon analysis of sequence variants of CBDAS and THCAS from different C. sativa L. strains, the CBDA synthase was considered as the ancestral synthase from which the THCAS evolved (Onofri et al., 2015)

Cytosol - Wikipedia

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Plastid - Wikipedia

en.wikipedia.org

Apoplast - Wikipedia

en.wikipedia.org

Natural gene variation in Cannabis sativa unveils a key region of cannabinoid synthase enzymes​

View ORCID ProfileCloé Villard, Christian Bayer, Nora Pasquali Medici, Arjen C. van de Peppel, View ORCID ProfileKatarina Cankar, Francel Verstappen, View ORCID ProfileIris F. Kappers, View ORCID ProfileM. Eric Schranz, View ORCID ProfileBastian Daniel, View ORCID ProfileRobin van Velzen
doi: https://doi.org/10.1101/2023.08.30.555511
Posted September 01, 2023.

https://www.biorxiv.org/content/10.1101/2023.08.30.555511v1.full.pdf

 

[acespicoli]

The total number of cannabinoid oxidocyclase genes varies considerably across cultivars. Onofri et al. (2015) amplified up to 5 (in cultivar “Haze”) different full-length fragments in chemotype I drug-type cultivars and up to 3 (in cultivars from Yunnan and Northern Russia and an inbred Afghan hashish landrace) different full-length fragments in chemotype III fiber-type cultivars. Inbred individuals of cultivars “Carmen” and “Skunk #1” are expected to be homozygous but yielded four and five cannabinoid synthase fragments, respectively (Weiblen et al. 2015). McKernan et al. (2015) detected up to six different fragments (including pseudogenes) of THCAS and related sequences. A recent study on copy number variation in cannabinoid oxidocyclase genes estimated that some of the analyzed cultivars could have up to 10 different fragments (Vergara et al. 2019). Based on these results it is clear that cannabinoid oxidocyclase genes can be considered a unique gene family that stems from a recent expansion and includes genes with unknown function (Onofri et al. 2015; Weiblen et al. 2015; Vergara et al. 2019; Hurgobin et al. 2021).

Recently, two drafts of the genome sequence of C. sativa L. have been published (www.medicinalgenomics.com; van Bakel et al., 2011), implemented by extensive transcriptome sequencing in different organs and strains. The publicly available database (http://genome.ccbr.utoronto.ca/) increased the number of known THCA- and CBDA-synthase gene sequences. It also became clear from this and other works (Kojoma et al., 2006) that there are many THCA- and CBDA-synthase-related pseudogenes in the Cannabis genome with several degrees of variation compared with the functional, chemotype-determining ones. Sequence variation was also observed within the putatively functional genes of both enzymes although, under the conditions in which the transcriptome was sequenced, the chemotypes expressed by the plants (the drug strain Purple Kush and the oil seed variety Finola), were not fully specified.

In the context of cannabis genetics, THCA synthase is generally considered dominant over its less active variants, meaning that even if a plant inherits one copy of a gene for high THCA synthase activity (dominant allele) and one copy for low activity (recessive allele), the plant will still produce high levels of THCA, the precursor to THC, due to the dominant allele's strong influence on the trait.

Key points about THCA synthase dominance:

• High THC strains:
  Plants with dominant THCA synthase alleles tend to produce high levels of THC, resulting in "drug-type" cannabis strains.
  
  
• Low THC strains:
  Plants with recessive THCA synthase alleles produce lower levels of THC, often categorized as "fiber-type" cannabis with lower psychoactive effects.
  
  
• Polymorphisms:
  Variations in the THCA synthase gene (polymorphisms) can lead to different levels of THC production, influencing the dominant or recessive nature of the trait in specific cannabis cultivars.
  

• Tetrahydrocannabinolic acid synthase - Wikipedia
  Polymorphisms of THCA synthase result in varying levels of THC in Cannabis plants, resulting in "drug-type" and "fiber-type" C. ..
  
  Wikipedia
  
  
  
• Dominance - Wikipedia
  In genetics, dominance is the phenomenon of one variant of a gene on a chromosome masking or overriding the effect of a different ...
  
  Wikipedia
  
  
  
• Tetrahydrocannabinolic acid - Wikipedia
  Tetrahydrocannabinolic acid (THCA, 2-COOH-THC; conjugate base tetrahydrocannabinolate) is a precursor of tetrahydrocannabinol (THC...
  
  
  

  SNP in Potentially Defunct Tetrahydrocannabinolic Acid ...​

  
  National Institutes of Health (NIH) (.gov)
  https://pubmed.ncbi.nlm.nih.gov › ...
  

by AR Garfinkel · 2021 · Cited by 28 — CBGA dominance is inherited as a single recessive gene, potentially governed by a non-functioning allelic variant of the THCA synthase.

 

[acespicoli]

 

[acespicoli]

The accumulation of Cu++ and Zn++ ions with zeatin induce feminization,
whereas the accumulation of Pb++ ions favors a masculinization effect

Front. Plant Sci., 03 November 2020
Sec. Plant Breeding
Volume 11 - 2020 | https://doi.org/10.3389/fpls.2020.569958

Genetic Architecture of Flowering Time and Sex Determination in Hemp (Cannabis sativa L.): A Genome-Wide Association Study​

Characteristic Reactions of Lead Ions (Pb²⁺)

Lead is a soft metal having little tensile strength, and it is the densest of the common metals excepting gold and mercury. It has a metallic luster when freshly cut but quickly acquires a dull color …

chem.libretexts.org

• Characteristic reactions of Pb2+Pb2+: The +2 oxidation state is the more stable state.

Index of genetics articles - Wikipedia

en.wikipedia.org

Plants (Basel)
. 2023 Jan 21;12(3):493. doi: 10.3390/plants12030493

The Cannabis Plant as a Complex System: Interrelationships between Cannabinoid Compositions, Morphological, Physiological and Phenological Traits​

 

[acespicoli]

Individuals possessing two BT alleles result in THC synthase (and THC) production. Individuals possessing two BD alleles result in CBD synthase (and CBD) production. Individuals with one BT and BD allele produce both THC and CBD.

Note: Selfed plants dramatically increase production levels

 

[acespicoli]

View full-text article in PMC
Plants (Basel)
. 2023 Nov 22;12(23):3927. doi: 10.3390/plants12233927

• Copyright and License information

Table 1.​

Evaluating natural triploids in 13 different Cannabis sativa genotypes.

Cultivar/Cross​	Source​	Plants Tested​	Number of Triploids​	Triploid Frequency (%)​
Bigfoot Glue​	Humboldt Seed Co., Eureka, CA, USA​	180​	1​	0.56​
Jelly Rancher​	Humboldt Seed Co., Eureka, CA, USA​	190​	0​	0.00​
DHN7 × DHN1​	Dark Heart, Davis, CA, USA​	1449​	7​	0.48​
Gazzurple​	Humboldt Seed Co., Eureka, CA, USA​	80​	0​	0.00​
DHN2 × DHN3​	Dark Heart, Davis, CA, USA​	263​	2​	0.76​
DHN2 × DHN4​	Dark Heart, Davis, CA, USA​	200​	1​	0.50​
DHN5 × DHN3​	Dark Heart, Davis, CA, USA​	200​	1​	0.50​
DHN6 × DHN3​	Dark Heart, Davis, CA, USA​	200​	1​	0.50​
DHN8 × DHN9​	Dark Heart, Davis, CA, USA​	1000​	3​	0.30​
DHN1 × DHN1​	Dark Heart, Davis, CA, USA​	128​	3​	2.34​
DHN7 × DHN1​	Dark Heart, Davis, CA, USA​	768​	2​	0.26​
DHN8 × DHN1​	Dark Heart, Davis, CA, USA​	337​	1​	0.30​
DHN10 × DHN1​	Dark Heart, Davis, CA, USA​	192​	0​	0.00​
Total​	​	5187​	22​	0.50 ± 0.603 *​

* This value represents mean ± standard deviation.

Front. Plant Sci., 29 April 2019
Sec. Plant Breeding
Pheno huntin >>> Volume 10 - 2019 | https://doi.org/10.3389/fpls.2019.00476

 

[acespicoli]

Tetraploid Phenotype​

Significant effects of ploidy were noted on plant growth and morphology. To generate material for this analysis, diploid and tetraploid strain 2 plants in tissue culture were transferred to soil and grown into mother plants. Fifteen cuttings per mother plant were rooted in soil for phenotypic assessment and chemical analysis.

The polyploid strain showed a reduction in rooting success. After 4 weeks, only 60% of tetraploid clones were successfully rooted (n = 9) compared to 100% of diploids (n = 15). Among rooted tetraploids, root emergence was slightly delayed (16.0 ± 3.7 days) compared to diploids (13.5 ± 4.7 days). Ploidy effects on leaf morphology were also observed. Tetraploids had larger fan leaves compared to diploids (Figures 3A,B). The central leaflet was significantly wider by an average of 0.75 cm on tetraploid leaves compared to diploid leaves, during the flowering phase (Figure 4A). Nail polish impressions showed that stomata on the underside tetraploid fan leaves were about 30% larger and half as dense compared to diploids (Table 2 and Figures 3C,D).


Figure 3

Figure 3. Leaf and stomata morphology. Representative images showing mature fan leaves of (A) diploid and (B) C. sativa strain 2 collected after 4 weeks of vegetative growth and 1 week under flowering lights. Scale bars, 2.5 cm. Nail polish impressions showing stomata on the abaxial surface of (C) diploid and (D) tetraploid fan leaves. Scale bars, 12 μm.


Figure 4

Figure 4. Growth parameters. Comparison of growth metrics in diploid (orange, n = 10) and tetraploid (blue, n = 9) C. sativa strain 2 plants. 5-week-old rooted clones were transplanted at week 0. Plants were moved to the flowering room at week 4 (arrowhead). Flowering lights were applied in week 5. (A) Width of the central leaflet in mature fan leaves. (B) Plant height from soil to highest meristem. (C) Diameter of the stem at 1 inch above the soil. (D) Sum of the length of all lateral stems. Data are means ± standard error. Asterisks indicate significant differences (Student’s t-test, p < 0.05).


Table 2

Table 2. Stomata size and density (mean ± SE) were measured on the abaxial side of mature fan leaves of diploid and tetraploid strain 2 C. sativa plants.


The height and stem base width of diploid and tetraploid plants were similar throughout growth. During the vegetative phase, tetraploid plants had slightly shorter lateral stems, but this difference was not significant following the switch to flowering (Figures 4B–D). Plants of both ploidies showed their first flowers after 1 week under flowering lights, and the rate of floral growth was similar throughout the flowering phase.

Trichome density on sugar leaves was measured at 2 weeks prior to harvest. Tetraploid leaves showed 40.4% higher glandular trichome density (4.41 ± 0.16 trichomes per mm2) compared to diploids (3.14 ± 0.15 trichomes per mm2). However, there was no obvious difference in the maturity of the trichomes on leaves, with the majority in the milky stage and some beginning to turn amber (Figure 5).


Figure 5

Figure 5. Trichome density. Representative images showing trichome density on the adaxial surface of the 4th sugar leaf of C. sativa strain 2 plants (A,B) diploid, (C,D) tetraploid. Leaves were imaged on the 7th week of flowering. Scale bars, 1 mm.


Front. Plant Sci., 29 April 2019
Sec. Plant Breeding
Pheno huntin >>> Volume 10 - 2019 | https://doi.org/10.3389/fpls.2019.00476

 

[acespicoli]

Trichome density on sugar leaves was measured at 2 weeks prior to harvest. Tetraploid leaves showed 40.4% higher glandular trichome density (4.41 ± 0.16 trichomes per mm2) compared to diploids (3.14 ± 0.15 trichomes per mm2). However, there was no obvious difference in the maturity of the trichomes on leaves, with the majority in the milky stage and some beginning to turn amber (Figure 5)

Pathologica
. 2023 Dec 1;115(6):302–307. doi: 10.32074/1591-951X-900

How to measure your microscope’s HPF. A critical guide for residents​

http://www.essentialpathology.info/gradingschema/index.html?HelpfulMethods

 

[acespicoli]

Cannabis is a Dioecious, Heterozygous, wind pollinated, and an Obligate Outcrosser, you need a minimum of 1000 Males and 1000 females all freely pollinating at the same time in the same location, otherwise you will lose genes every reproduction, this is what is required for conservation, breeding is different, requiring more or less plants, dependant on the goals.
@Sam_Skunkman

Critical Reviews in Plant Sciences · November 2016 DOI: 10.1080/07352689.2016.1267498
Was nice for a friend to point these factors out so long ago

 

[acespicoli]

Population genetics - Wikipedia

en.wikipedia.org

Landrace - Wikipedia

en.wikipedia.org

Inbred strain - Wikipedia

en.wikipedia.org

Polyploidy - Wikipedia

en.wikipedia.org

Cloning - Wikipedia

en.wikipedia.org

Genes 2021, 12(6), 923; https://doi.org/10.3390/genes12060923

 

[acespicoli]

Dark Horse Genetics​

Here you can find all information about the cannabis breeder Dark Horse Genetics. We've collected data about 70 Cannabis Strains breed by Dark Horse Genetics (1 of this strains got reviews of the SeedFinder users, with an average rating from 7.33 out of 10!) Click on the strains to find more informations, pictures, reviews, comparisons and sources for a variety - and/or check out the Breeder Info here at the page to find out more about Dark Horse Genetics.

Dark Horse Genetics's strains​

Strain name	Breeder	Flowering time	Strain heritage	Feminized
303 Stooges	Dark Horse Genetics	70	mostly indica	regular
808 Mana	Dark Horse Genetics	63	unknown	regular
Banners Revenge	Dark Horse Genetics	70	mostly sativa	regular
Bitch Slap	Dark Horse Genetics	65	mostly indica	regular
Bloody Zundae	Dark Horse Genetics	67	indica / sativa	regular
Brazilian Bombshell	Dark Horse Genetics	-	mostly indica	regular
Bruce Banner	Dark Horse Genetics	63	mostly indica	reg. and fem.
Bruce Banner 1.0	Dark Horse Genetics	70	mostly indica	regular
Chem Berry D	Dark Horse Genetics	67	mostly sativa	regular
Chem Jong-Un	Dark Horse Genetics	63	indica / sativa	regular
Chem Lem	Dark Horse Genetics	-	indica / sativa	regular
Cherry Wonka	Dark Horse Genetics	70	unknown	regular
Clown Shoes OG	Dark Horse Genetics	70	unknown	regular
Conjugal Visit	Dark Horse Genetics	63	indica / sativa	regular
Cry Baby	Dark Horse Genetics	70	indica / sativa	regular
Dark Gamma Kush	Dark Horse Genetics	70	mostly indica	regular
Dark Heart	Dark Horse Genetics	70	unknown	regular
Dark Horse OG	Dark Horse Genetics	70	mostly indica	regular
Double Lemon Pie	Dark Horse Genetics	63	mostly sativa	regular
Face Melt OG	Dark Horse Genetics	70	unknown	regular
Flash Glue	Dark Horse Genetics	60	mostly indica	regular
Galactus	Dark Horse Genetics	-	mostly indica	regular
Gamma Berry	Dark Horse Genetics	67	indica / sativa	regular
Grape Cobbler	Dark Horse Genetics	63	unknown	regular
Grape Smash	Dark Horse Genetics	63	indica / sativa	regular
Hulk Smash	Dark Horse Genetics	67	indica / sativa	regular
Hulkamania	Dark Horse Genetics	63	indica / sativa	regular
Ice Cream Zundae	Dark Horse Genetics	63	indica / sativa	regular
Joe's Lemonade F2	Dark Horse Genetics	63	mostly sativa	regular
Kings Banner XIII	Dark Horse Genetics	70	mostly indica	regular
Kings Bounty	Dark Horse Genetics	70	unknown	regular
L.A. Sunshine	Dark Horse Genetics	70	unknown	regular
Lazar 115	Dark Horse Genetics	63	indica / sativa	regular
Lemon Cream	Dark Horse Genetics	56	mostly sativa	regular
Lemon Head	Dark Horse Genetics	56	mostly sativa	regular
Lemon Jedi OG	Dark Horse Genetics	60	mostly indica	regular
Lemon Twizzler	Dark Horse Genetics	-	indica / sativa	regular
Meyer Lemons	Dark Horse Genetics	60	mostly indica	regular
Mind Glazer	Dark Horse Genetics	70	unknown	regular
Mind Ztone	Dark Horse Genetics	63	unknown	regular
Mizzizzippi Mud	Dark Horse Genetics	63	indica / sativa	regular
Mr. Zoftee	Dark Horse Genetics	67	indica / sativa	regular
Orange Blossom Fizz	Dark Horse Genetics	70	mostly sativa	regular
Orange Cream	Dark Horse Genetics	70	mostly sativa	regular
Original Gorilla Glue #4	Dark Horse Genetics	63	mostly sativa	feminized
Phantom Juliet	Dark Horse Genetics	56	unknown	regular
Power Ztone	Dark Horse Genetics	63	unknown	regular
Reality Ztone	Dark Horse Genetics	63	unknown	regular
Roid Rage	Dark Horse Genetics	63	indica / sativa	regular
Runthz	Dark Horse Genetics	63	indica / sativa	regular
Sagerbloom Haze	Dark Horse Genetics	67	mostly sativa	regular
Savage Hulk	Dark Horse Genetics	63	indica / sativa	regular
Simbiote	Dark Horse Genetics	63	indica / sativa	regular
Solomon Grundy	Dark Horse Genetics	70	mostly indica	regular
Soul Ztone	Dark Horse Genetics	63	indica	regular
Sour Lemons	Dark Horse Genetics	63	unknown	regular
Sour ThanoZ	Dark Horse Genetics	63	unknown	regular
Space Ztone	Dark Horse Genetics	63	unknown	regular
Stockton Slap	Dark Horse Genetics	63	indica / sativa	regular
Strawberry Diesel	Dark Horse Genetics	63	mostly sativa	regular
Strawberry Glue	Dark Horse Genetics	67	mostly indica	regular
Strawberry Shortcake	Dark Horse Genetics	69	mostly sativa	regular
Sunny D	Dark Horse Genetics	70	indica / sativa	regular
Super Slutty Haze	Dark Horse Genetics	70	mostly sativa	regular
Suzy Q OG	Dark Horse Genetics	70	unknown	regular
ThanoZ	Dark Horse Genetics	56	indica / sativa	regular
Tiki Drink	Dark Horse Genetics	60	mostly sativa	regular
Time Ztone	Dark Horse Genetics	63	unknown	regular
Vitamin C	Dark Horse Genetics	65	indica / sativa	regular
Weapon X	Dark Horse Genetics	63	indica / sativa	regular

Dark Heart

Landos "wide" leaf male - leaf mass and stomata density

Imagine folks discarding plants based solely on wld vs nld

 

[acespicoli]

https://hemp.cals.cornell.edu/resources/reports-factsheets/2022-cornell-hemp-field-day-handouts/triploid-pollen-challenge-trials/

 

[acespicoli]

Fig. 7.
(A) Example leaflets for CBD dominant cultivars, (B) intermediate cultivars, and (C) THC dominant cultivars. (D) Example mature inflorescences of cultivars with green leaves and green calyx, (E) green leaves with purple calyx, and (F) purple leaves with purple calyx. (G) Example of compact inflorescences, (H) loose inflorescences, and (I) inflorescences infected with Botrytis cinerea.

Citation: HortScience horts 56, 4; 10.21273/HORTSCI15607-20

Hope you find the latest interesting and most of all beneficial in your breeding projects
Best >>>

 

[Mithridate]

Im clinically retarded.

This has nothing to do with the following, just wanted to get it out there.

I went down a "leaf morphogenesis" rabbit hole recently. Weed is an infinitely maleable plant where most cues about morphology mean something just not what we (at least I?) would presume.

This blew my mind. I didn't register shat and will revisit the subject shortly. Ha!

 

[acespicoli]

Phenotype Plasticity
@Mithridate im full of theory's, its great when I can find a white paper to agree

Observer effect (physics) - Wikipedia

en.wikipedia.org

 

[Mithridate]

The mechanics remain unchanged. The double slit experiment proved one thing and one thing only, its that through ego we force our limited comprehension on complex systems and blurt out silly exceptions to rules we know nothing about.

Leaf angle from blade to blade, width, length, serrations, ribs etc are all.. gulp.. hormonal, bacterial and nutritional

 

[Mithridate]

Echoes of a 4th dimension
No direct correlation to plants here but
Are you familiar with Walter Russell? Tesla told his friend Walter to hide his books for 1000 years for humans are not ready to appreciate.

Cool dude

 

[acespicoli]

Walter Russell - Wikipedia

en.wikipedia.org

List of Nikola Tesla writings - Wikipedia

en.wikipedia.org

Wish I had more time for reading, very satisfying past time
Would be nice to convert to audio files

 

[acespicoli]

An autosome is any chromosome that is not a sex chromosome.[1] The members of an autosome pair in a diploid cell have the same morphology, unlike those in allosomal (sex chromosome) pairs, which may have different structures.
https://en.wikipedia.org/wiki/Autosome

Plant genera Cannabis and Humulus share the same pair of well-differentiated sex chromosomes First published: 12 May 2021 https://doi.org/10.1111/nph.17456​

Discussion​

We here identified the Humulus lupulus sex chromosomes, and found that they are homologous to those of Cannabis sativa (Prentout et al., 2020), and that a part of these chromosomes had already stopped recombining in a common ancestor of the two species. Performing a segregation analysis with SEX-DETector (Muyle et al., 2016), we identified 265 XY genes in H. lupulus, among which 112 also were sex-linked in C. sativa. Mapping these genes on the chromosome-level assembly of C. sativa (Grassa et al., 2018) suggested that the nonrecombining region is large in H. lupulus, as proposed before based on cytological studies (Divashuk et al., 2011).

We identified three different regions on the sex chromosome, based on the distribution of sex-linked gene phylogenetic topologies and synonymous divergence between the X and Y copies on the C. sativa X chromosome: one region that had already stopped recombining in a common ancestor of C. sativa and H. lupulus, a region that independently stopped recombining in both species, and the pseudo-autosomal region. Our results suggest the pseudo-autosomal boundary (PAB) in H. lupulus may be located around 20 Mb, whereas we estimated a PAB around 30 Mb in C. sativa (Prentout et al., 2020); the nonrecombining region thus may be larger in H. lupulus than in C. sativa. With this estimation of the size of the nonrecombining region in H. lupulus, among the 3469 genes present on the X chromosome, 2045 genes would be located in this nonrecombining region (which represents 59.1% of all the genes on the X chromosome). However, a chromosome-level assembly of the H. lupulus genome would be needed to determine the exact position of the PAB in this species, as synteny might not be fully conserved. In addition, because we used one single cross, it is possible that we overestimated the size of the nonrecombining region as a consequence of linkage disequilibrium. Thus, genes around the PAB classified as sex-linked and for which we estimated a low dS value may still be recombining. An accurate estimation of the PAB, as has been done for example in Silene latifolia, would require much more offspring and data from several populations (Krasovec et al., 2020).

Several sex-linked genes had topologies that were not compatible with either recombination suppression in a common ancestor or in each of the species independently. Strikingly, most of these topologies placed the H. lupulus Y sequence as an outgroup to the other sex-linked gene sequences. Whether this is the result of errors (e.g. long-branch attraction, mapping biases) remains to be investigated. Interestingly, genes with theses ‘unexpected’ topologies all clustered (except for one gene) in a region of c. 25 Mb. This region is located at the extremity of the X chromosome, which, as we suggested, stopped recombining first. It is likely to observe a high rate of unexpected phylogenetic results in the region that stopped recombining first because the X–Y divergence should be the highest in this region, which could increase the mapping bias. Our approach to correct for the Y read mapping relies on geneconv, which is known to have a high rate of false negatives (Lawson & Zhang, 2009). This also could explain the unexpected presence of some of the XY-XY genes in the older region.

X–Y gene conversion has been shown to affect only a few genes in animals (Katsura et al., 2012; Trombetta et al., 2014; Peneder et al., 2017). Although we do not expect gene conversion for half of the genes that are sex-linked in both species, it is worth noting that a part of fragments identified by geneconv may correspond to real gene conversion rather than mapping biases. Here again, assemblies of Y and X chromosomes in both species are required to determine the presence of true X–Y gene conversion.

The highest dS values and the genes with a topology indicating that recombination was already suppressed in the common ancestor are located in the same region (65 Mb to the end of the chromosome). These results suggest the presence of at least two strata in these sex chromosomes. We estimated that the youngest stratum is 10.1–29.4 Myr old in H. lupulus, and 15.9–19.8 Myr old in C. sativa (Table S5). However, although recombination suppression clearly did not occur for all of the sex-linked genes at the same time, we cannot determine the exact number of strata in the sex chromosomes of C. sativa and H. lupulus. It also is possible that recombination was suppressed gradually, with the recombination suppression starting before the split of both genera and continuing afterwards. To clearly determine the number of strata, an identification of chromosomal inversions or significative differences in dS values along the sex chromosomes is required (Nicolas et al., 2004; Lemaitre et al., 2009; Wang et al., 2012; reviewed in Wright et al., 2016). Thus, X and Y chromosome assemblies for both H. lupulus and C. sativa are needed to exactly determine the number (and location) of strata in both species. Moreover, a Y chromosome assembly will allow the identification of Y-specific genes, which is not possible with SEX-DETector and the data that we used.

We did not find X-hemizygous genes in H. lupulus. This is striking as 218 X-hemizygous genes (38% of all sex-linked genes) were found in C. sativa using the same methodology (Prentout et al., 2020). A very low level of polymorphism could result in the inability of SEX-DETector to identify X-hemizygous genes (Muyle et al., 2016), but in that case SEX-DETector also should have problems identifying autosomal genes, which was not the case here. Nonrandom X-inactivation in females could be an explanation, as the nonrandom expression of a single X allele in females would impede SEX-DETector to identify X-linkage and X-hemizygous genes (Muyle et al., 2016). We ran an Allele-Specific Expression (ASE) analysis, which did not support this hypothesis (Figs S5–S7). Humulus lupulus probably is an ancient polyploid that reverted to the ancestral karyotype (Padgitt-Cobb et al., 2019). It is thus possible that the H. lupulus X chromosome comprises two copies of the ancestral X as some cytological data seem to suggest (Divashuk et al., 2011). In this case, SEX-DETector would manage to identify the XY gene pairs, but would fail to identify the X-hemizygous genes as these genes would exhibit unexpected allele transmission patterns (Fig. S8).

Humulus lupulus is a rare case of XY systems in plants in which the Y is smaller than the X (cf. Ming et al., 2011). In C. sativa, both sex chromosomes have similar sizes (Divashuk et al., 2014). If the size difference is caused by deletions of parts of the H. lupulus Y chromosome, which is the hypothesized mechanism in many species (cf. Ming et al., 2011), we expect to observe that many XY gene pairs in C. sativa have missing Y copies in H. lupulus. As explained above, we did not detect any X-hemizygous genes. Furthermore, the XY gene pairs of H. lupulus were distributed uniformly on the C. sativa X chromosome, and no region appeared to be depleted in XY genes, which is not what we would observe if large deletions were present on the H. lupulus Y chromosome. The sex chromosome size differences observed in H. lupulus probably reflect complex dynamics, different from that of old animal systems with tiny Y chromosome resulting from large deletions (e.g. Skaletsky et al., 2003; Ross et al., 2005). The large size of the X chromosome in H. lupulus may be due to a full-chromosome duplication followed by a fusion (see above), whereas the Y chromosome has remained unchanged. Assemblies of the H. lupulus sex chromosomes will be needed to test these hypotheses.

Our estimates of the age of the H. lupulus sex chromosomes are larger than the estimates for C. sativa, although we found very similar X–Y maximum divergence in both species (higher bound age estimates are c. 50 Myr and c. 28 Myr old; highest dS values are 0.362 and 0.415 in H. lupulus and C. sativa, respectively, see Prentout et al., 2020). Of course, the molecular clocks that we used are known to provide very rough estimates as they derive from the relatively distant Arabidopsis genus, and are sensitive to potential differences in mutation rates between the annual C. sativa and the perennial H. lupulus (Neve, 1991; Petit & Hampe, 2006; Small, 2015; but see Krasovec et al., 2018). Indeed, only one of these molecular clocks (which is based on the mutation rate) takes into account the generation time (2 yr in H. lupulus vs 1 yr in C. sativa). This produced age estimate is approximately two-fold greater than that from the other clock (based on the substitution rate), which was not the case with C. sativa (Prentout et al., 2020). It is not known, however, if the generation time influences the substitution rate (Petit & Hampe, 2006). Furthermore, the short generation time in C. sativa probably is a derived trait, not reflecting the long-term generation time of the Cannabis–Humulus lineage, as the Cannabis genus is the only herbaceous genus in the Cannabaceae family (Yang et al., 2013). Thus, the remarkable similarity between the highest dS values in both species indicates that the C. sativa and H. lupulus sex chromosomes have a similar age, as expected if they derive from the same common ancestor. Although it is not possible to estimate their age exactly with the current data, initial recombination suppression at least pre-dates the split between the genera, that occurred between 21 and 25 Myr ago (Ma) (Divashuk et al., 2014; Jin et al., 2020), and might even be 50 Myr old. We thus confirmed here that the XY system shared by C. sativa and H. lupulus is among the oldest plant sex chromosome systems documented so far (Prentout et al., 2020).

Dioecy was inferred as the ancestral sexual system for the Cannabaceae, Urticaceae and Moraceae (Zhang et al., 2019; note, however, that many monoecious Cannabaceae were not included). We found that the synonymous divergence between the Cannabaceae species and Morus notabilis was approximately 0.45, higher than the maximum divergence of the X and Y copies in the Cannabaceae. It remains possible that the sex chromosomes evolved before the split of the Cannabaceae and Moraceae families, because the oldest genes might have been lost or were not detected in our transcriptome data. There is, however, no report of whether or not sex chromosomes exist in Urticaceae and Moraceae (Ming et al., 2011).

In order to estimate the Y expression, we counted the number of reads with Y SNPs. Therefore, the impact of a potential Y reads mapping bias should be weaker on Y expression analysis than on X–Y divergence analysis. We validated this assumption by removing genes with detected mapping bias from the analysis, which did not change the signal of Y expression reduction and dosage compensation (Table S6; Figs S9, S10). Dosage compensation is a well-known phenomenon in animals (e.g. Gu & Walters, 2017), but it has been documented only quite recently in plants (reviewed in Muyle et al., 2017). Here we found evidence for dosage compensation in H. lupulus; this is not surprising as previous work reported dosage compensation in C. sativa and we showed here that both systems are homologous. Cannabis sativa and H. lupulus add up to the list of plant sex chromosome systems with dosage compensation (see Muyle et al., 2017, for a review; and Prentout et al., 2020, and Fruchard et al., 2020, for the latest reports of dosage compensation in plants). Further analyses are needed to determine whether this dosage compensation has been selected or is an outcome of regulatory feedback (Malone et al., 2012; Krasovec et al., 2019).

Humulus lupulus sex chromosomes, like those of C. sativa, are well-differentiated, with a large nonrecombining region. Both species show similar patterns of Y degeneration and dosage compensation, despite the fact that a large part of the nonrecombining region evolved independently in both species. These similarities, as well as the age of the chromosomes and the fact that they have been conserved since the most recent common ancestor of the two genera – a unique situation in plants so far – provide an exciting opportunity to test and elaborate hypotheses on sex chromosome evolution in plants.

 

[acespicoli]

Sounds unrealistic. The % in the link add up to 113.8%

I submitted a email to them pointing out the issue with the data @chilliwilli hopefully they can find the cause and correct it. nice catch !