High concentrations of sucrose induce overwintering bud formation in gentian plantlets cultured in vitro

Abstract

Overwintering buds (OWBs) are dormant organs produced by perennial plants. After flowering, Japanese gentians (Gentiana triflora, G. scabra and their hybrids) produce OWBs in autumn and go dormant to survive winter. Although dormant OWBs have been reported to have strong cold and freezing tolerances, no details about mechanisms and regulation are known. We attempted to produce in vitro OWBs (IOWBs) from gentian plantlets to facilitate the study of OWBs. We found that high-sucrose (6%) culturing media induced IOWB formation. IOWBs showed an enhanced freezing tolerance together with high levels of proteins and transcripts know to specifically accumulate in OWBs. Furthermore, IOWBs showed endodormancy release and bud break in the same manner as OWBs. Bud breaking in OWBs and IOWBs was promoted by red light and inhibited by far-red light. These results indicated that IOWBs possess similar physiological characteristics as OWBs, at least regarding stress tolerance and dormancy. Therefore, our artificial IOWB induction technique will be useful for the study of physiological mechanisms in field-grown OWBs.

A novel virus transmitted through pollination causes ring-spot disease on gentian (Gentiana triflora) ovaries

“…Bands corresponding in molecular mass to the GORV coat protein were excised and digested with trypsin (Promega). The tryptic peptides were desalted with SPE C-TIP (AMR) and subjected to nano liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis as described previously (Imamura et al, 2014). Peptides were identified using a MASCOT MS/MS ion search in error tolerance mode using a private database containing the genomic cDNA sequences of GORV.…”

The Gentio-Oligosaccharide Gentiobiose Functions in the Modulation of Bud Dormancy in the Herbaceous PerennialGentiana

“…These OWBs were harvested from plants grown in an agricultural field at the Iwate Biotechnology Research Center from September 2009 to April 2010 or September 2010 to April 2011, frozen in liquid nitrogen, and lyophilized. G. triflora cv Ihatovo was used for in vitro culture, and IOWBs were induced by the method described by Imamura et al (2014).…”

“…Since IOWBs had dormancy characteristics similar to field-grown OWBs (Imamura et al, 2014) and the sugar contents of IOWBs were easily controlled ( Figure 6A; Supplemental Figure 5), IOWBs can be used for studying the effects of sugars on budbreak. IOWBs were cultured on solid sugar-free Murashige and Skoog (MS) medium or MS medium containing 1% (w/v) gentiobiose for 3 weeks.…”

“…Quantification of gene transcript levels by quantitative RT-PCR analysis was conducted according to the method of Imamura et al (2014). For synthesizing first-strand cDNA, 500 ng of total RNA was reverse transcribed using a High Capacity cDNA reverse transcription kit (Applied Biosystems) with random primers according to the manufacturer's instructions.…”