Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress

Verona, Italy - 24/27 September, 2003

ISBN 88-900622-4-X

Poster Abstract - 3.03

CANNABIS SATIVA: A GENETIC PLANT MODEL FOR INDUSTRIAL END USES

P. RANALLI, G. MANDOLINO, A. CARBONI, M. DI CANDILO

Istituto Sperimentale per le Colture Industriali, Via di Corticella 133, 40128 Bologna

multi-use crop, cannabinoids, sexual differentiation, industrial uses

New crops are required for specialist needs, including raw materials for the chemical and pharmaceutical industries (bioethanol from sugar crops, fuel from oil crops, energy from biomass production). As a result, the redeployment of surplus agricultural resources and a switch away from dependence on fossil resources have been attempted. The development of plants for specialised productivity needs an enhanced understanding of metabolic control both in terms of molecular and genetic mechanisms and the consequences of remodelling of metabolic pathways. Hemp is a multi-use crop able to provide fiber, cellulose, seeds and seed oil, cannabinoids and biomass. Textile, paper, composite, fuel, energy, and pharmaceutical products are developed from this raw material. Integrating many agroindustrial chains, Cannabis is considered a crop model in which insights into specific metabolic pathways and biosynthetic processes are valuable for improvement of the plant for all sets of industrial derivatives. The research on hemp, supported by a project funded by the Italian Ministry of Agriculture, dealt with the following tasks: 1) the recovering and maintenance of hemp germplasm accessions in a viable condition and their reproduction; new methods of hemp genetic resource evaluation based on DNA analysis were also performed; 2) the identification of molecular markers for relevant traits (male phenotype and THC genes) and their allocation in a saturated linkage map were performed. The identification of QTL linked to other important characters, such as fibre content, is an important task in progress. In detail, efforts were focused to gain insight into the genetic factors underlying the sexual differentiation and the expression of the most important cannabinoid categories stored in the plant. Cannabis is a dioecious species with unisexual flowers and sex determined by heteromorphic chromosomes (X and Y). The gene expression of male and female apices was compared by cDNA AFLP. Seven of the several polymorphic fragments have been identified to be differentially expressed which belong to nine different mRNAs induced to female apices. The results of BLAST analysis for these clones led to the identification of their putative coding function and to their possible roles in the earliest step of sexual differentiation. Furthermore, the two most relevant cannabinoids synthesized in Cannabis plants are delta-9-tetrahydrocannabinol (delta⁹THC, psycoactive) and cannabidiol (CBD, non psychoactive) both originated from the common precursor cannabigerol (CBG, non psychoactive) by two enzymes THC-and CBD-synthase. Genetic analysis of inbred lines, and of progenies F1 and F2 support a monogenic model controlling the chemotype: one single locus (B) with two codominant alleles, BD and BT, each probably encoding one of the two synthases involved in cannabinoid biosynthesis; the F2 generations fitted the expected 1:2:1 chemotype segregation ratio. Sequence-specific markers for each of the two alleles involved in chemotype determination were also developed. Another relevant target is to increase the seed oil content and composition of cannabis oil. To date, cloning of lipid-associated genes from Cannabis have not been reported. However, the availability of heterologous genes for direct introduction into hemp, or their availability to design primers for isolating the corresponding cannabis homologous by PCR-based techniques is relatively advanced. Finally, fiber improvement is assisted by metabolic pathway engineering accomplished in cotton for introducing modified alcaligene eutrophus genes encoding the enzymes acetacetyl-CoA reductase and polyhydroxyalkanoate synthase. These two enzymes, when produced in fibre cells along with endogenous B-ketothiolase, catalyse the polymerization of acetyl-CoA to polyhydroxybutyrate. The transgenic fibres exhibited better insulating properties. The isolation of other relevant genes for the remodelling of pathways is a diminishing hurdle thanks to the pace of DNA sequence analysis of small plant genomes and the proliferation of EST (expressed sequenced tag) libraries. The huge genetic diversity exhibited by hemp germplasm allows access to relevant genes for specific functions. The advances in hemp basic and applied research are elucidated in this review; there are elements to identify strategy for design modified plants with enhanced industrial end uses.