Factors Affecting Wheat(Triticum aestivum L.)Transformation Mediated by Biolistic Bombardment

The plasmid pDB1 with gus-bar as double marker gene,and the plamsid pARN6、pBAB3、pYAO24 with RC24 gene encoding rice basic chitinase were co-transferred to immature embryos of 8 wheat genotypes in combination of pDB1+pARN6,pDB1+pBAB3 or pYAO24 via particle bombardment.pYAO24 has nptⅡ as selective gene.Transgenic plants were regenerated after selective culture.Factors influencing wheat transformation via particle bombardment,such as genotype,gold particle preparation,parameters of bombardment,were analyzed.It is found that Xinong 88 is a very ideal genotype of genetic transformation,immature embryos two weeks post-pollination were suitable wheat materials of bombardment,30-50 μg gold particle per bombardment was appropriate,mixing the gold particle suspension thoroughly could reduce the injury to immature embryos and enhance the transformation frequency.GUS histochemical staining revealed that blue spots produced by sucrose as osmoticum were bigger than that of mannitol as osmoticum.150 mg/L kanamycin was adopted when nptⅡ was used as selective gene.It can produce false positive plants easily that the phosphinothricin(PPT) concentration was 2 mg/L when bar was used as selective gene.We should enhance the concentration form 2 mg/L to 3-5 mg/L in the future.