用全DNA转化法构建多功能石油降解菌

CONSTRUCTION OF MULTI FUNCTIONAL OIL DEGRADATIVE BACTERIA BY TOTAL DNA TRANSFORMATION

以嗜蜡的不动杆菌４－１ 作为受体菌，经溶菌酶破壁后，用嗜胶的假单胞菌Ｚ１７ 的全ＤＮＡ 转化，经再生后用只含高胶油的选择培养基筛选出转化子不动杆菌ＺＨ２０．破壁及再生的最佳条件为：溶菌酶浓度１．５ｍ ｇ／ｍ Ｌ，３７℃水浴保温４０ｍ ｉｎ，破壁率为９３．８％ ，再生率达到３０．２％ ；转化条件为：３０％ ＰＥＧ６０００，５０ｍ ｍ ｏｌ／ＬＣａＣｌ２ ，于３７℃保温２０ｍ ｉｎ．ＺＨ２０ 具有嗜高胶油和高蜡油的双重性状，５０℃条件下使高胶油粘度降低２２．２％ ，使高蜡油的凝固点降低２．５℃，且保持了受体菌的耐热和耐盐性能．液蜡培养基连续传代１０ 次，性能稳定．

DNA extracted from Pseudomonas sp. strain Z 17 , which is an asphalt philic strain, was transformed to Acinetobacter sp. strain 4 1, a paraffin philic strain under the condition of 30% PEG6000, 50mmol/L CaCl 2, incubated at 37℃ for 20min. After regeneration, a transformant Acinetobacter sp. strain ZH 20 was selected on the plate which contained the only carbon resource of asphalt. This transformant can emulsify both high asphalt oil and high paraffin oil. At 50℃, it can decrease the viscosity of high asphalt oil by 22.2%, and can decrease the solidifying point of high paraffin oil by 2.5℃. At the same time, the strain ZH 20 keeps the characteristic ability of high temperature tolerance and salt tolerance. After growing for 10 generations in liquid paraffin media, the oil degradative functions of the strain ZH 20 were proved to be stable.