Obtaining New Lines of Wheat Cells with RNAi-mediated Resistance against Plant-Parasitic Nematodes using New Microbial Bioregulators (original) (raw)

2018, 2nd International Conference on Innovations in Natural Science and Engineering, 07-10 September. 2018, Kyiv, Ukraine

Impact of new microbial bioregulators: Avercom, Avercom nova-2, Violar and Phytovit on induction of callus formation and regeneration of plantlets of wheat (Triticum aestivum L.) of cultivar Zimoyarka on the MS nutrient media was studied under in vitro conditions. The increase of frequency of formation of callus cells (from 43 up to 74 %) was observed on the apical meristem of 4-5-day-old seedlings of wheat placed on the MS media supplemented with microbial bioregulators used at the concentrations ranging from 10 up to 60 μl/l in combination with 1 mg/l auxin 2.4-D as compared with lower frequency of formation of callus cells obtained on the MS media supplemented with only auxin 2.4-D used at the concentration 1 mg/l (control). The increase of efficiency of regeneration of wheat shoots (from 35 up to 65 %) was observed on the 3-4-week-old callus tissue grown on the MS media supplemented with microbial bioregulators used at the concentrations ranging from 20 up to 100 μl/l in combination with 1-2 mg/l cytokinin BAP and 0.5-1 mg/l auxin IAA as compared with lower efficiency of regeneration of wheat shoots obtained on the MS media supplemented with 1-2 mg/l cytokinin BAP and 0.5-1 mg/l auxin IAA (control). The rooting of shoots was shown on the MS media supplemented with microbial bioregulators used at the concentrations ranging from 10 up to 100 μl/l. We conducted also physiological and molecular-genetic studies of resistance of wheat plants obtained under in vitro conditions to plant parasitic nematodes. In vitro experiments showed that wheat plants grown on MS media with microbial bioregulators had 20-37 % of infestation with cereal cyst nematode Heterodera avenae, significantly lower when compared with 73 % of infestation of the control plants grown without bioregulators. Bioregulators increased morphometric parameters of the wheat plants obtained under in vitro conditions and further grown under greenhouse conditions on the natural invasive background: stem height increased 13.6-37.5 %, flag leaf length 18.0-19.3 %, ear length 6.8-24.6 %, and ear weight 27.0-54.5 %, when compared with control wheat plants. The difference in the degree of hybridized molecules mRNA and small regulatory si/miRNA from control and experimental wheat plants increased: 15-39 % in plants grown under in vitro conditions on the invasive background created by H. avenae and 33-56 % in seeds of plants grown under greenhouse conditions on the natural invasive background. The silencing activity of si/miRNA from wheat plants grown on MS media with bioregulators increased: 20-51 % in plants grown under in vitro conditions on the invasive background created by H. avenae or 38-64 % in plants grown under greenhouse conditions on the natural invasive background. Our studies confirm the RNAi-mediated resistance to plant nematodes of wheat plants grown under in vitro and greenhouse conditions. Keywords: MS media, phytohormones, microbial bioregulators, wheat callusogenesis and organogenesis, plant resistance to parasitic nematodes, small regulatory si/miRNA.