TITLE:
Efficient Technique for Long-Term in Vitro Storage of Transgenic Aspen Genotypes
AUTHORS:
Elena O. Vidyagina, Konstantin A. Shestibratov
KEYWORDS:
Populus tremula, Long-Term Storage, In Vitro Culture, Osmolytics, Transgenic Genotypes, Microshoots
JOURNAL NAME:
American Journal of Plant Sciences,
Vol.9 No.13,
December
13,
2018
ABSTRACT: In vitro culture of isolated cells from tissues and organs
is sometimes used to preserve and reproduce unique genotypes of woody plants.
The technique, however, requires regular subculturing which raises storage
costs and creates risks for contamination and accumulation of somaclonal variations.
We examined the effects of sugar composition of culture medium, the length of
photoperiod, light intensity, and ambient temperature on the survival of plant
material in vitro. The study was performed on 49 genotypes of Populus
tremula (46
transgenic genotypes carrying GFP-, Xeg- and Gus-genes, and 3 control (wild-type) genotypes). It was
shown that effective storage of plants was achieved through optimization of the
combined effects of all storage parameters under study. Based on the
experimental data, we developed a protocol for long-term in
vitro storage of desirable genotypes without subculture and with a survival rate of
up to 98%. The best results were obtained when the plant material was
pre-cultured on a WPM medium containing 15 g/L sucrose, 7.5 g/L sorbitol and
7.5 g/L mannitol, and then stored at +4°C under
a 24-hour light day cycle with only 8 hours of light per day and maximum light
intensity of 2000 lux. Post-storage recovery was done by culturing on a medium
containing 1 mg/L gibberellic acid. The developed method can be used for
effective in vitro storage of the studied genotypes for up to 24
months without subculture.