农杆菌介导的BADH—CMO基因转化玉米愈伤组织的初步研究

Preliminary Studies on Transforming Bivalent Salt-Tolerant Gene into Maize Callus by Agrobacterium tumefaciens

植物的耐盐性是一个多基因控制的复杂性状,将多个耐盐相关基因导入同一植物更有助于提高植物的耐盐能力。以玉米自交系H99、丹988的胚性愈伤组织为受体材料,通过愈伤组织对NaCl的敏感性试验,确定转化愈伤组织的适宜选择压,适宜愈伤组织转化的盐浓度为225 mmol/L。并对农杆菌遗传转化系统的条件进行了初步研究,用带有pCAMBIA-1301-CMO-BADH质粒的根癌农杆菌转化玉米愈伤组织,结果表明:采用EHA105的菌株振荡20 h后,菌液OD值为0.5～0.6时侵染25 min为农杆菌转化的适宜条件。对移栽成活的36棵抗盐植株经PCR检测有2株呈阳性,阳性株率最高达到5.6%,初步证明目的基因已整合到玉米基因组中。

Salt tolerance of plants is a complex trait controlled by multiple genes and the transformation of multiple genes into the same salt-tolerant plants helps to improve salt tolerance of plants.Embryogenic calli of inbred H99 and Dan 988 were selected.Using NaCl sensitivity experiments,suitable callus selection pressure was determined,and salt concentration 225 mmol/L was most suitable for callus tumefaciens.The conditions of Agrobacterium tumefaciens transforming system were studied.Anti-sense expression vector of pCAMBIA-1301-CMO-BADH gene was transformed into maize inbred callus by Agrobacterium tumefaciens.The results show that： EHA105 strain vibrated 20 h,concentration of fermenting liquor was OD 0.5—0.6 and infection time was 25 min.2 plants among 36 plants surviving selection by PCR amplification were positive,transformation ratio reached 5.3%.The preliminary studies show that the genes have been transferred into the genome of maize successfully.