农杆菌介导的假俭草遗传转化体系的建立

摘要/Abstract

摘要： 以假俭草优良种质‘E126’为受体材料,以HPT基因为报告基因,通过农杆菌介导法转入ZjDREB1基因。研究了转化体系中筛选剂和抑菌素对胚性愈伤组织生长和绿苗分化的影响,以及菌液浓度、侵染时间和共培养时间对转化效率的影响。结果表明,愈伤筛选和再生苗筛选的最佳潮霉素浓度均为30 mg/L,头孢霉素作为抑菌素的最佳浓度为400 mg/L;菌液OD₆₀₀值为0.3,侵染30 min,共培养3 d为最优转化条件。经PCR检测,初步获得10株转基因植株。

Abstract: Agrobacterium tumefaciens strain LBA4404 harboring the plasmid pCAMBIA1301 which contains stress tolerance-related transcription factor ZjDREB1 gene and reporter gene HPT was used to transform axillary bud-derived embryogenic calluses of Eremochloa ophiuroides ‘E126’. To systematically optimize the conditions of centipedegrass transformation, several factors which could influence Agrobacterium-mediated DNA transfer were examined, including concentration of selective agent and bacteriostat, bacterial concentration, duration of infection and co-cultivation. Consequently, a transformation system of transgenic centipedegrass plants was established. It was found that the efficiency of transformation highly relied on the following optimal conditions: The concentration of hygromycin for selection was 30 mg/L; the cefotaxime concentration for restraining the Agrobacterium was 400 mg/L; OD₆₀₀ as 0.3, 30 min of infection time and 3 days. Co-cultivation time was the optimized condition for genetic transformation system in E. ophiuroides. Ten transgenic plants were identified by PCR detection from initial 44 regenerated plants.

中图分类号:

张芳,王舟,宗俊勤,刘建秀,佘建明. 农杆菌介导的假俭草遗传转化体系的建立[J]. 草业学报, 2011, 20(2): 184-192.

ZHANG Fang, WANG Zhou, ZONG Jun-qin, LIU Jian-xiu, SHE Jian-ming. Establishment of an Agrobacterium mediated genetic transformation system for Eremochloa ophiuroides[J]. Acta Prataculturae Sinica, 2011, 20(2): 184-192.

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