Double haploid development in drug-type Cannabis sativa L. through microspore indirect de-novo plant regeneration

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Abstract

Double haploid technology (DH) is an essential tool in plant breeding, enabling the rapid production of homozygous lines. However, Cannabis sativa L. has been categorized as recalcitrant to DH induction. In this study, we evaluated the potential to generate DH C. sativa plants via anther culture and indirect de-novo organogenesis. We examined two cultivars, a THCA-dominant cultivar and a CBDA-dominant line. Callus induction success varied, with 29.48% for the THCA cultivar and 71.08% for the CBDA genotype with a regeneration success of 14.45% in 17 weeks for the latter. Mixoploidy in the callus indicated spontaneous genome doubling, while genetic testing confirmed DH nature of the regenerants. This is the first report documenting the successful induction of DH C. sativa plants through de-novo indirect organogenesis. These findings have profound implications for the C. sativa breeding sector by potentially improving efficiency of genome editing and hybrid development in this economically significant species.

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