Author/Authors :
-، - نويسنده Department of Plant Breeding, Faculty of Agriculture, Tarbiat Modares University, P.O. Box 14115-336, Tehran, I.R. Iran Gharanjik, Shahrokh , -، - نويسنده Department of Plant Breeding, Faculty of Agriculture, Tarbiat Modares University, P.O. Box 14115-336, Tehran, I.R. Iran Moieni, Ahmad , -، - نويسنده National Institute of Genetic Engineering and Biotechnology, P.O. Box 14965/161, Tehran, I.R. Iran Mousavi, Amir , -، - نويسنده Department of Plant Breeding, Faculty of Agriculture, University of Tehran, P.O. Box 4111, Karaj, I.R. Iran Alizadeh, Houshang
Abstract :
Haploid microspore-derived embryos (MDEs) of wheat were obtained by in vitro androgenesis. These embryos were employed to evaluate the transient expression of GUS gene (uidA) following particle bombardment. Using the Bio-Rad PDS-1000/He system, the physical parameters including rupture disk pressure (900, 1100 and 1350 psi); microprojectile travel distance (6 and 9 cm); gold particles size (0.6 mm, 1 mm and 1.6 mm), DNA and microcarrier concentrations (0.5 mg of DNA with 150 μg of gold particles or 1.0 μg of DNA with 300 mg of gold particles/bombardment) and bombardment numbers (1x (single) and 2x (double)) were assessed. The effect of high osmoticum in the bombardment medium (0.3 M mannitol and 0.4 M maltose) and the age of embryos were also evaluated. Optimal expression in MDEs was obtained using the following conditions of double bombardment at 1350 psi, 9 cm target distance, a 1 mm gold particle size, 1.0 mg of DNA with 300 mg of gold particles/bombardment, and osmotic pretreatment of 4-6 weeks old embryos using 0.4 M maltose for 6 h before and 16 h after bombardment. The optimized transformation protocol presented in this study is expected to improve devalopment of commercial transgenic wheat lines expressing desirable agronomic traits.
