Production of Tetraploid and Triploid Hemp

Abstract

To maximize yield, cannabidiol (CBD) hemp producers prefer female plants, and this is accomplished by using expensive feminized seed, vegetatively propagated female clones, or by removing male plants from dioecious seed lots. Hemp pollen drifts long distances on wind, and pollination of females reduces CBD content. Induction of triploidy is a common strategy used by plant breeders to produce sterile cultivars of agricultural crops. Triploid (3n) hemp, with three sets of chromosomes, was developed by crossing naturally diploid (2n) hemp with tetraploid (4n) hemp. Tetraploid plants used to create triploids were produced using pregerminated seeds and the mitotic spindle inhibitor colchicine. Seedlings from seeds of ‘Abacas’ × [(‘Otto2’ × ‘BaOx’) × (‘BaOx’ × ‘Colorado Cherry’)] treated with 0.05% colchicine or 0.02% colchicine for 12 hours and longer were significantly shorter than controls and ≤1 cm tall at 10 days after sowing. Surviving seedlings exhibited thickened cotyledons and hypocotyls, which indicated a potential change in ploidy. Tetraploid induction ranged from 26% to 64% for pregerminated seeds of five different hemp cultivars (Abacus × Wife, Cherry Wine, Mountain Mango, Wife, and Youngsim10) treated with 0.05% colchicine for 12 hours. Tetraploids had nearly twice the DNA content as diploids according to flow cytometric analysis. Tetraploid ‘Wife’ had larger stomates and reduced stomatal density compared with diploid ‘Wife’. Four triploid ‘Wife’ genotypes produced from crossing tetraploid ‘Wife’ with diploid ‘Wife’ were acclimated to greenhouse conditions after embryo rescue. DNA content and stomate size of triploid ‘Wife’ was intermediate between the parents. This is the first report of triploid plants of hemp. Future research will evaluate the sterility of triploid hemp.

Characteristics of the Diploid, Triploid, and Tetraploid Versions of a Cannabigerol-Dominant F1 Hybrid Industrial Hemp Cultivar, Cannabis sativa ‘Stem Cell CBG’

“…Based on the reduction in seed development as compared to other crossing combinations, the triploids were determined to be infertile. Infertile hemp has been recommended as a strategy to mitigate yield reduction caused by pollination [ 13 ]. As far as we are aware, this research represents the first report of triploid C. sativa female infertility.…”

“…[ 9 ], Humulus lupulus [ 10 ], Hypericum androsaemum [ 11 ], and Miscanthus sinensis [ 12 ]. Reports have speculated that triploid C. sativa cultivars may be infertile, and therefore resistant to the yield damage caused by pollination [ 13 ]; however, we were unable to locate studies demonstrating triploid hemp sterility.…”

“…Several ploidy manipulations in C. sativa have been reported in the literature [ 13 , 23 , 24 ]; however, knowledge of the effects on chemical profiles is limited to the differences between tetraploid and diploid type I and type II plants, those that are tetrahydrocannabinol (THC)-dominant, or those that produce both THC and cannabidiol (CBD), respectively. These studies indicated that although THC concentration remained unchanged in polyploids, CBD production was slightly increased [ 23 , 24 ].…”

“…These studies indicated that although THC concentration remained unchanged in polyploids, CBD production was slightly increased [ 23 , 24 ]. A method of producing triploid type III plants, those that are CBD-dominant, has also been reported [ 13 ]; however, the field phenotype and fertility status of the triploid plants were not discussed. Furthermore, we were unable to locate any studies discussing the impact of ploidy manipulation on type IV, cannabigerol (CBG)-dominant, cultivars.…”

Advances and Perspectives in Tissue Culture and Genetic Engineering of Cannabis

“…Cannabis is naturally a diploid and multiple successful artificial inductions of polyploidy in cannabis have been reported [ 159 , 160 , 161 ]. Kurtz et al [ 160 ] have used this technique to produce triploid plants, but field performance and lack of seed development have not yet been reported. These reports provide promising results for the potential for polyploidy to be used to improve cannabis cultivars.…”