A novel and rapid method for Agrobacterium-mediated production of stably transformed Cannabis sativa L. plants

Galán-Ávila, Alberto; Gramazio, Pietro; Ron, Mily; Prohens, Jaime; Herraiz, Francisco Javier

doi:10.1016/j.indcrop.2021.113691

Cited by 26 publications

(36 citation statements)

References 45 publications

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“…We believe that 5th generation energy crops could be even more improved through somaclonal variation and/or genetic manipulation to obtain a sample with the lowest amount of lignin. Since clustered regularly interspaced short palindromic repeats (CRISPR)-mediated genome editing and Agrobacterium -mediated gene transformation have been developed in cannabis, genetic manipulation of callus can be considered a promising tool to change the lignin pathways in order to produce more bioenergy 29 – 31 . This study suggests a new approach for future research projects that utilize genetically modified crops for biofuel production.…”

Section: Resultsmentioning

confidence: 99%

In vitro plant tissue culture as the fifth generation of bioenergy

Norouzi

Hesami

Pepe

et al. 2022

Sci Rep

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Developing and applying a novel and sustainable energy crop is essential to reach an efficient and economically feasible technology for bioenergy production. In this study, plant tissue culture, also referred to as in vitro culture, is introduced as one of the most promising and environmentally friendly methods for the sustainable supply of biofuels. The current study investigates the potential of in vitro -grown industrial hemp calli obtained from leaf, root, and stem explants as a new generation of energy crop. For this purpose, the in vitro grown explants were first fully characterized in terms of elemental and chemical composition. Secondly, HTL experiments were designed by Design Expert 11 with a particular focus on biocrude. Finally, the chemical components, functional groups, and petroleum-like hydrocarbons present in the biocrude were identified by PY-GCMS. A 22.61 wt.% biocrude was produced for the sample grown through callogenesis of the leaf (CL). The obtained biocrude for CL consisted of 19.55% acids, 0.42% N compounds, 15.44% ketones, 16.03% aldehydes, 2.21% furans, 20.01% aromatics, 5.2% alcohols, and 19.88% hydrocarbons. To the best of the authors’ knowledge, this is the first report that in vitro -grown biomass is hydrothermally liquefied toward biocrude production; the current work paves the way for integrating plant tissue culture and thermochemical processes for the generation of biofuels and value-added chemicals.

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Section: Resultsmentioning

confidence: 99%

In vitro plant tissue culture as the fifth generation of bioenergy

Norouzi

Hesami

Pepe

et al. 2022

Sci Rep

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“…We undertook further work to examine if there is a causal relationship between CsMIXTA expression and trichome development in a heterologous system. Cannabis is recalcitrant to genetic transformation; a few reports have been published recently [ 16 , 17 , 18 ], but their transformation efficiencies are not robust. In lieu of a reliable cannabis transformation protocol, we used a heterologous system to determine if there is a causal effect of CsMIXTA expression on trichome morphogenesis.…”

Section: Resultsmentioning

confidence: 99%

Overexpression of CsMIXTA, a Transcription Factor from Cannabis sativa, Increases Glandular Trichome Density in Tobacco Leaves

Haiden

Apicella

et al. 2022

Plants

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Cannabinoids are synthesized in glandular stalked trichomes on the female flowers of Cannabis sativa (cannabis). The regulation of glandular trichome development has not been characterized in cannabis. We recently identified an R2R3-MYB transcription factor, CsMIXTA, which could be involved in trichome morphogenesis in cannabis. Some homologous genes of CsMIXTA are known to function in glandular trichome initiation in other plant species. CsMIXTA is highly expressed in flower tissue compared to vegetative tissues. Interestingly, CsMIXTA is also highly expressed in trichomes isolated from female flower tissue. In addition, CsMIXTA is upregulated during the peak stages of female flower maturation in correlation with some cannabinoid biosynthetic genes. Transient expression in Nicotiana benthamiana showed that CsMIXTA is localized in the nucleus. Furthermore, yeast transcriptional activation assay demonstrated that CsMIXTA has transactivation activity. Overexpression of CsMIXTA in Nicotiana tabacum resulted in higher trichome density, larger trichome size, and more branching on stalked glandular trichomes. The results indicate that CsMIXTA not only promotes glandular trichome initiation in epidermal cells, but also regulates trichome development in tobacco leaves. In this report, we characterized the novel function of the first cannabis transcription factor that may be critical for glandular trichome morphogenesis.

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“…Some regenerants had an albino phenotype on the 100 mg/L kanamycin selection medium but spontaneous in vitro regeneration and rooting were evident in certain transformed lines. This study by Galán-Ávila et al [101] also indicated that the Futura75 strain had the highest transformed hypocotyl-derived transgenics that were transiently expressing the GUS reporter gene, whereas for the FINOLA, Fedora17 and Felina52 varieties, no transgenics were produced, further emphasising on the genotype-dependent specificity aspect that is integral to biotechnological manipulations of Cannabis.…”

Section: Genetic Engineering and Gene Editingmentioning

confidence: 89%

“…Attempts to use Agrobacterium-mediated transformation are ongoing as researchers try to expand the varieties that are being tested with this method. Concomitantly to assessing the effectiveness of agrobacterial co-cultivations with seedlings of the short-day C. sativa agricultural lines (i.e., Ferimon, Felina32, Fedora17, USO31 and Futura71) and the day-neutral FINOLA type with the intention of generating transgenic plants, an in vitro regeneration protocol was assessed when plants were placed on 0.2 mg/L NAA: 4 mg/L TDZ MS medium [101]. Although cotyledons, hypocotyls and apical meristem explants were co-incubated with Agrobacterium tumefaciens LBA4404 (pBIN19), as expected, not all these explants had similar transformation rates.…”

Section: Genetic Engineering and Gene Editingmentioning

confidence: 99%

Cannabis sativa: From Therapeutic Uses to Micropropagation and Beyond

Adams

Malatsi

Makunga

2021

Plants

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The development of a protocol for the large-scale production of Cannabis and its variants with little to no somaclonal variation or disease for pharmaceutical and for other industrial use has been an emerging area of research. A limited number of protocols have been developed around the world, obtained through a detailed literature search using web-based database searches, e.g., Scopus, Web of Science (WoS) and Google Scholar. This article reviews the advances made in relation to Cannabis tissue culture and micropropagation, such as explant choice and decontamination of explants, direct and indirect organogenesis, rooting, acclimatisation and a few aspects of genetic engineering. Since Cannabis micropropagation systems are fairly new fields, combinations of plant growth regulator experiments are needed to gain insight into the development of direct and indirect organogenesis protocols that are able to undergo the acclimation stage and maintain healthy plants desirable to the Cannabis industry. A post-culture analysis of Cannabis phytochemistry after the acclimatisation stage is lacking in a majority of the reviewed studies, and for in vitro propagation protocols to be accepted by the pharmaceutical industries, phytochemical and possibly pharmacological research need to be undertaken in order to ascertain the integrity of the generated plant material. It is rather difficult to obtain industrially acceptable micropropagation regimes as recalcitrance to the regeneration of in vitro cultured plants remains a major concern and this impedes progress in the application of genetic modification technologies and gene editing tools to be used routinely for the improvement of Cannabis genotypes that are used in various industries globally. In the future, with more reliable plant tissue culture-based propagation that generates true-to-type plants that have known genetic and metabolomic integrity, the use of genetic engineering systems including “omics” technologies such as next-generation sequencing and fast-evolving gene editing tools could be implemented to speed up the identification of novel genes and mechanisms involved in the biosynthesis of Cannabis phytochemicals for large-scale production.

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A novel and rapid method for Agrobacterium-mediated production of stably transformed Cannabis sativa L. plants

Cited by 26 publications

References 45 publications

In vitro plant tissue culture as the fifth generation of bioenergy

In vitro plant tissue culture as the fifth generation of bioenergy

Overexpression of CsMIXTA, a Transcription Factor from Cannabis sativa, Increases Glandular Trichome Density in Tobacco Leaves

Cannabis sativa: From Therapeutic Uses to Micropropagation and Beyond

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