Using orthogonal design to determine optimal conditions for intergeneric protoplasts fusion between Mingxian169 and Y2155a

Abstract

This study is important for the optimization of protoplast fusogen and washing solution system suitable for protoplast fusion between the Triticum aestivum and Aegilops. By enzymolysis, the result shows that more than 90% viable protoplasts of Mingxian169 (common wheat) and Y2155a (Aegilops) were efficiently obtained and fused. The greatest of protoplast fusogen and washing solution condition was developed using an orthogonal experimental design, L25 (55), where L=orthogonal table; 5=factors; 5=five levels of each; and 25=experimental number. It was shown that the greatest protoplast fusogen was found at PEG6000 content (w/v) 25%, 0.50 M sucrose, pH 6.0 and washing solution was 5.0 mM CaCl₂. Over 10% viable heterokaryons was produced using different fusion condition. Because of the narrow genetic diversity of common wheat and elite agronomic traits of many wild relatives, it is very important and helpful for the improvement of common wheat through somatic hybridization between wheat and wheat wild relatives,

Key words: Protoplast, fusion, orthogonal design method, Mingxian 169, Y2155a.