, 2010). In the same study, it was observed that the HSP30p-mediated expression of FLO11 ORF in either BM45 or VIN13 did not generate a flocculent phenotype under either standard laboratory media or synthetic MS300 must fermentation conditions. In the present study, we demonstrate that HSP30p-FLO11-based transgenic BM45 and VIN13 wine yeast strains are capable of a novel

MI-flocculation phenotype that seems to exclusively occur under authentic red wine fermentation conditions. This flocculation phenotype selleckchem can be characterized as being partially Ca2+ dependent and Ca2+ independent. In particular, we show that HSP30p-FLO11 transgenic wine yeast strains displaying this trait were able to produce significantly clearer wines with compacted buy Z-VAD-FMK lees fractions. All yeast strains used in this study are listed in Table 1. Yeast strains were routinely cultivated at 30 °C in rich YEPD medium, containing 1% yeast

extract, 2% peptone and 2% glucose. For selection of sulphometuron methyl (SM)-resistant BM45 and VIN13 transformants, SC medium containing 0.67% YNB and 2% glucose was supplemented with 280 and 300 μg mL−1 SM (DuPont Agricultural Products, France), respectively. Yeast strains were cryopreserved in YEPD supplemented with 15% glycerol (Ausubel et al., 1995). The cell density of suitably diluted yeast cell suspensions in 100 mM EDTA was manually determined using a haemocytometer. Grapes of Vitis vinifera Merlot (200 kg) were rinsed with sulphited water, destemmed and crushed.

As a precaution, damaged grape clusters (broken or with visual microbial alterations) were discarded in order to eliminate undesirable contamination. Red grape must [24.2% sugar (glucose and fructose), 5.8 g L−1 titratable acidity and pH 5.8] was sulphited to 40 mg L−1. Evodiamine Thereafter, red grape must was batch fermented in 20-L plastic buckets containing 3 kg of Merlot grape juice that was adjusted to exactly 10 kg by the addition of a mixture consisting of grape pulp and skins. This was followed by the addition of 4 g of diammonium phosphate. Yeast precultures in YEPD were prepared and processed as described previously (Govender et al., 2008). Thereafter, wild-type and transgenic yeast inoculum populations were preacclimatized for wine fermentations by incubation at 30 °C for 4 h with shaking at 160 r.p.m. in filter (0.22 μm cellulose acetate)-sterilized 50% v/v Merlot juice diluted with distilled water. The fermentative potential of BM45 and VIN13 wild-type strains and their transgenic derivatives were assessed in triplicate. Assuming a ratio of 0.