Improved in ovulo embryo culture for stenospermocarpic grapes (Vitis vinifera L.)

Abstract

In ovulo embryo rescue techniques have been used to recover new hybrids from seedless × seedless grape crosses. This study was conducted to increase efficiency by investigating effects of genotype, medium, and ovule removal age on ovule elongation, embryo recovery, growth, and plantlet formation. Ovules from self-pollinated berries of seedless varieties Sunmuscat, Merbein Seedless, and Marroo Seedless were cultured at 30, 43, 60, and 70 days after flowering (DAF) in a range of media, some of which were supplemented with gibberellic acid (GA3) and indole-3-acetic acid (IAA). The effect of activated charcoal (AC) in media on rescued embryos was also investigated. Ovules exhibited continuous growth in vivo and in vitro. The most vigorous growth was observed for ovules cultured at 30 and 43 DAF, but more embryos were recovered from ovules cultured at 60 and 70 DAF. Ovule growth and embryo production in vitro were improved in Bouquet and Davis (BD) and Nitsch and Nitsch (NN) media. Supplementation with GA3 increased embryo recovery rates. Highest embryo recovery rates were 18.1%, 9.6%, and 12.2% for Sunmuscat, Merbein Seedless, and Marroo Seedless, respectively, when ovules were excised and cultured at 60 or 70 DAF in either BD or NN media. In vitro embryo survival and plantlet formation were higher for torpedo-shaped embryos, and improved greatly in 6-benzyladenine (BA)-supplemented woody plant (WP) medium containing 0.3% AC. Embryo recovery was improved by excising and culturing ovules at 60 DAF in BD or NN media and then by transferring embryos to WP medium supplemented with BA and AC.