麦草畏降解菌株的分离、鉴定及降解特性研究

发布时间：2018-03-27 19:48

  本文选题：麦草畏　切入点：微生物降解　出处：《南京农业大学》2015年硕士论文

【摘要】：麦草畏在我国被广泛的运用在农业除草方面,它主要被作为出苗后除草剂使用,用来防治多年生阔叶杂草和禾本科杂草。因为它的结构较为稳定和过度的使用,导致它在土壤中的残留十分严重,所以越来越多人开始研究麦草畏在土壤中的降解问题。国外已经有能降解麦草畏的菌株被分离得到的报道,也有相关降解途径和基因的报道,但在国内研究的比较少。本研究从麦草畏降解菌株的分离与筛选开始,进而研究其理化特性及降解特性、推测相关的降解途径并且克隆降解的基因。利用麦草畏为唯一碳源,利用富集驯化技术从农药厂废水生化处理污泥中分离到具有麦草畏降解能力的菌株,命名为SC-1,根据它的培养特征、生理生化特征和16s rDNA分析可以将其初步鉴定为鞘酯菌属。该菌对麦草畏的降解效率可达到90%,它的最适生长温度为30℃,最适的pH为7.0左右,不耐盐;对于无机氮源的利用率较低,而对葡萄糖、麦芽糖和糊精等糖源的利用率却比较高。SC-1可以为麦草畏为唯一的的碳源生长,且在36 h内几乎可以将1g/L的麦草畏降解完全,在温度为30℃、Na+浓度为10 g/L、pH为7左右时,菌株的降解效率较好。其降解时的最佳环境与SC-1生长所需要的环境很相似,同样也是在接种量和通气量较大的情况下降解的效果比较好。当麦草畏的浓度低于1000mg/L时,SC-1的对其的降解效率都比较好,当麦草畏浓度高于1000mg/L后,会对SC-1产生毒性,SC-1不但无法降解麦草畏甚至无法生长。利用液相色谱和质谱对SC-1降解麦草畏的中间产物检测结果表明,该菌降解麦草畏的第一步为脱甲基生成没有除草剂活性的中间产物3,6-DCSA(3,6--二氯水杨酸)。将麦草畏降解为DCSA的这一步中起关键作用的基因是DMO基因,该基因主要调控麦草畏的O-型脱甲基酶合成,该酶主要有还原酶、铁氧化还原蛋白和加氧酶组成。此酶系统主要用作于电子传递链,在电子传递链中NADH(烟酰胺的还原形式腺嘌呤二核苷酸)穿梭通过一个还原酶到铁氧还蛋白。国外对此的研究比较多,关于DMO的报道也比较的多,所以有关DMO的基因序列已经报道过了,根据已经报道的基因序列设计引物后,将SC-1的DNA作为模板扩增后进行测序,发现其序列与已报道的相同,所以基本可以确定SC-1对麦草畏的降解途径与已报道的完全相同。
[Abstract]:Wheat straw forceps are widely used in agricultural herbicides in China. They are mainly used as herbicides after emergence to control perennial broadleaved weeds and gramineous weeds because of their stable structure and excessive use. As a result of its very serious residue in the soil, more and more people began to study the degradation of wheat straw forceps in the soil. There have been reports of the isolation of strains that can degrade the forbidding wheat straw, as well as related degradation pathways and genes. But there are few studies in our country. This study began with the isolation and screening of strains of wheat straw forvos degradation, and then studied its physical and chemical properties and degradation characteristics, speculated the related degradation pathway and cloned the degradation gene. By means of enrichment and acclimation, a strain with biodegradable ability of wheat straw was isolated from the sludge of biochemical treatment of agricultural pharmaceutical plant wastewater, named SC-1, according to its culture characteristics, Physiological and biochemical characteristics and 16s rDNA analysis showed that the strain could be identified as sphingomycetes. The biodegradation efficiency of the strain could reach 90%, the optimum growth temperature was 30 鈩，

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