The effect of supplementing in vitro flax (Linum usitatissimum L.) culture media with a synthetic cytokinin derivative (BAP9 THP) and a cytokinin metabolism modulator (INCYDE)

Abstract Main research objective of this work was to evaluate the potential of novel synthetic cytokinin (CK) derivatives and modulators of their metabolism in stimulating shoot bud organogenesis (multiple shoot formation) across various in vitro cultures of flax/linseed ( Linum usitatissimum L.) hypocotyls; as this developmental process is key in clonal micropropagation, the quality of newly propagated shoots was also assessed. The efficiencies of the CK derivatives used in this research were compared to what was observed for a frequently used aromatic synthetic CK, namely, 6-benzylaminopurine. In addition, the levels of endogenous isoprenoid and aromatic CKs in cultured hypocotyls were measured and followed to study potential interactions between endogenous and exogenous CKs, especially in terms of possible effects on regeneration ability. The experiments involved three fiber flax cultivars and one linseed cultivar. In general, both the synthetic derivatives of CKs and modulators of CK metabolism (BAP9THP, INCYDE) increased the frequency of multiple shoot organogenesis from hypocotyl segments, as well as resulted in the desired morphology for further practical utilization (number of buds and shoots, shoot lenght, negligible callus formation), relative to endogenous CKs. Endogenous CKs (IPR, IPR5´MP, cZROG) were present at low concentrations in the cultured explants, and – in some cases – completely absent (tZ, cZ, IP, IPG9); aromaticCKs were present at higher levels when compared to isoprenoid CKs, with BAP and BAPR5´MP proving to be the most abundant in the samples. All four of the detected aromatic CKs (BAP, BAPR, BAP9G, BAPR5´MP) showed the highest levels in plants grown on medium L-T6 supplemented with BAP and synthetic CK BAP9THP at a concentration ratio of 0.05:1. Impact of interactions between the endogenous and exogenous CKs on regeneration capacity was analyzed by a comparison of treatments. In addition to a CK-specific effect, a genotype-mediated effect on regeneration potential was detected during the experiments The linseed cultivar AGT583/05 exhibited the best characteristics across all recorded traits and all the tested media when compared to the three fiber flax cultivars (Venica, Jitka, Tábor). The obtained results will contribute to a better understanding of the conditions needed for flax regeneration and micropropagation.