Cellular responses of leaf explants of Cocos nucifera L. in vitro

Abstract

Leaf explants of Cocos nucifera L. (coconut palm) were studied in vitro in order to establish whether or not rapid cellular changes contribute to the well known recalcitrance of coconut cells in tissue culture. Segments from the base of immature leaves were cultured on modified Eeuwens' medium at 30°C in darkness. The mitotic index, nuclear DNA amounts, cell and nuclear size were measured both before and during culture (from 0 to 70 days). There was no basipetal gradient of cell division in immature coconut leaves; the mitotic index never exceeded 2% and showed neither a positional nor temporal relationship with leaf development. Moreover the vast majority of cells were in G1 of the cell cycle. This cell cycle pattern was maintained for most of the period in culture although at 70 days there was an increase in the proportion of cells in S-and G2-phases consistent with low rates of callus formation. The nuclear : cell size ratio was constant in cells within the immature leaf irrespective of developmental age. However upon transfer to culture media, cell size but not nuclear size increased. We suggest that this uncoupling of cell and nuclear size disrupts cell co-ordination and is a key contributor to recalcitrant cellular behaviour of this species in vitro.

Characterisation of a cyclin-dependent kinase (CDKA) gene expressed during somatic embryogenesis of coconut palm

“…The analyzed CDKA sequences differed mainly at the extreme 3 0 region however, generally confirmed the claim of Joubès et al (2000) indicating the CDKA family contains a highly conserved amino acid sequence with 89% similarity. In vitro culture reduces the tissue division rate (Winkelmann et al 1998), which holds true in coconut tissues (Jesty and Francis 1992;Sandoval et al 2003). In vitro cultured coconut explants have a high proportion of cells in the G0/G1 phase (*90%; Sandoval et al 2003), highlighting the slowness of the morphogenesis process during in vitro regeneration of coconut palm: in most explants, calli develop 3-4 months after inoculation on the callogenesis medium (Buffard-Morel et al 1992;Verdeil et al 1994, Chan et al 1998Pérez-Nuñez et al 2006).…”

“…Coconut tissues that have been used as explants for in vitro culture exhibit a low mitotic index. The immature leaves contained less than 1% of actively dividing cells (Jesty and Francis 1992). Furthermore, a progressive slowing of the growth was determined during in vitro culture in the G0/G1 phase by flow cytometric analysis (Sandoval et al 2003).…”

Genome-wide identification and characterization of the cyclin gene family in Populus trichocarpa

“…The M1-cyclins, including the A-and B-cyclins, assist in driving cells into the M-phase. The G1-cyclins, such as cyclins C, D, and E, among others, become active towards the end of the G1-phase and are responsible for ushering the cell into the S-phase (Jesty and Francis 1992).…”

The cell cycle in plant development

“…Again, generalizations about the level of polyploidy exhibited by an explant in culture and its ability to regenerate are fraught with difliculty. Some species seem able to tolerate mixoploidy in culture and continue to exhibit regeneration whilst others are highly recalcitrant yet primary cultures do not exhibit mixoploidy (Cocos nucifera, Jesty & Francis, 1991).…”