We have established a successful protocol for the genetic transformation of cannabis (C.
sativa L.). More stages of research were performed to establish the protocol for
adventitious regeneration of cannabis. We have first tested the different regenerative
potentials of three explant types (petioles, leaf cuttings, and internodes), and their
potential to produce callus. The effect of the DKW (Driver in Kuniyuki, 1984), and MS
(Murashige in Skoog, 1962) basic media was also explored. The influence of the light
regime on the regenerative potential of petioles, and the organogenetic potential of 6
medicinal genotypes one industrial cultivar of cannabis was also the subject of research.
The different concentrations of phytohormones thidiazurone (TDZ), 6-
benzylaminopurine (BAP), and 1-naphtaleneacetic acid (NAA), were also investigated
to find the most promising combination for callus production and shoot development.
Petioles from the BF.KEM1.2 genotype, showed the highest potential for callus and
shoots development compared with the other two explant types. Adventitious
regeneration of non-transformed shoots was also successfully established, with a 5%
rate of success, from the petioles of the BF.KEM3 genotype, on the DKW medium
supplemented with 0,2 mg/l of TDZ, 0,5 mg/l of NAA, and 0,1 μM/l of the
phytosulfokine (PSK) peptide. The presence of light positively influenced the shoots
and callus development. Furtherly, petioles from the genotype BF.KEM3 were
transformed by co-cultivation with Agrobacterium tumefaciens, which was carrying the
pCAMBIA1302-ZsGreen plasmid, which contains in its T-DNA region selection gene
hptII and reporter gene ZsGreen. We have confirmed the presence of the ZsGreen
reporter gene in 95 % of the examined transformed calli and the presence of the hptII
gene was confirmed in 85 % of the examined transformed calli.
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