不同系统pJP4质粒介导基因强化降解2,4-D效应研究

以携带pJP4质粒的基因工程菌Pseudomonas putida SM1443∷gfp2x(pJP4∷dsRed)为供体菌，考察了pJP4质粒在4种纯菌中的转移效应；并分别针对活性污泥、生物膜、颗粒污泥和河流沉积物系统，通过实验室小试实验考察了该基因工程菌对不同系统目标污染物2,4-二氯苯氧乙酸（2,4-D）的强化降解效应.结果表明，该基因工程菌中的pJP4质粒能够以广泛的微生物细胞为受体菌发生水平转移；向活性污泥、生物膜、颗粒污泥和河流沉积物系统加入一定量的基因工程菌，对2,4-D的降解都能够产生明显的促进作用.对于活性污泥系统（2,4-D初始浓度为450 mg/L），强化与对照系统反应143.5　h时2,4-D去除率分别为66％和54％；对于生物膜系统（2,4-D初始浓度为180　mg/L），强化与对照系统在反应113　h时对其去除率分别为99％和61％；对于颗粒污泥系统（2,4-D初始浓度为160　mg/L），强化系统2,4-D在62　h接近完全去除，而对照系统66　h去除率仅为26％；对于沉积物系统（2,4-D初始浓度为2　mg/L），强化与对照系统344　h去除率分别为93％和69％.激光共聚集扫描显微（CLSM）分析揭示并证实了不同基因强化系统接合子的形成与存在.

Plasmid pJP4 Mediated Gene-augmentation in Different Systems and Its Effect on 2,4-D Biodegradation

With pJP4 plasmid carrying genetic engineering bacteria Pseudomonas putida SM1443 :: gfp2x ( pJP4:: dsRed) as the donor, horizontal gene transfer of pJP4 plasmid in 4 isolated pure strains was investigated, and effects of the donor bacteria inoculation on the removal of the target pollutant 2,4-dichlorophenoxyacetic acid (2,4-D) was studied through conducting gene augmentation in activated sludge, biofilm, aerobic granular sludge and river sediment system, respectively. Results showed that plasmid pJP4 could transfer from Pseudamonas putida SM1443 to a broad spectrum of bacteria. Inoculation of pJP4 plasmid carrying donor bacterium apparently promoted the degradation of 2,4-D for all the above four systems. For the activated sludge system (2,4-D initial concentration at 450 mg/L) , 66% and 54% removal of 2,4-D was achieved after 143.5 h reaction for the gene augmented and control system, respectively. For the biofilm system with 2,4-D initial concentration at 180 mg/L, 2,4-D removal percentage at 113 h was 99% and 61% , respectively. For aerobic granular sludge system(2,4-D initial concentration at 160 mg/L), 2,4-D was nearly completely removed by 62 h in the gene-augmented system, while the control system only degraded 26% at 66 h. For the system with sediment (2,4-D initial concentration at 2 mg/L), 93% and 69% removal of 2,4-D was obtained at 344 h reaction for the gene augmented and control system, respectively. Confocal laser scanning microscopy (CLSM) analysis revealed the formation and presence of transconjugants in different gene augmentation systems.