Optimization of effective factors on β-glucuronidase (GUS) gene transformation by Agrobacterium to Trachyspermum ammi

Trachyspermum ammi is one of the most important medicinal herbs in Iran. This plant contains important medicinal properties, including anti-digestive, antispasmodic and relieving rheumatic pains and has been used as a spice and food preservative. To overexpress some effective ingredients in this plant, it is necessary to optimize the gene transfer process. For this purpose, the explants of hypocotyl, two strains of Agrobacterium tumefaciens were studied along with factors such an inoculation times (5, 10, 15, 20 min) co-cultivation times (1, 2 and 3 days). In this study, pBI121 vector harboring GUS as the reporter gene, and nptII gene as the plant selected marker were used. To confirm the gene transfer, PCR, histochemical GUS staining and RT-PCR tests were used. According to the results, the time of inoculation of 5 minute and the time of one day for co- cultivation increased the efficiency of gene transfer. Both bacterial strains showed a relatively good performance in gene transfer, but the LBA4404 strain showed higher efficacy than GV3101. In this study, 25 mg/L of kanamycin was selected as the appropriate concentration for determining the probable transgenic plant by Agrobacterium. For the first time, the present study presents an effective and efficient method for gene transfer to Trachyspermu ammi that can be effectively used for gene transfer.