一种产邻苯二酚菌株高通量筛选方法

A Method of High Throughput Screen of Catechol-Producing Strain

目的:对产邻苯二酚菌株进行筛选。方法:采用前期筛得的产邻苯二酚菌为出发菌株,通过硫酸二乙酯诱变的方法使该菌株突变,同时建立96孔板培养和酶标仪检测方法对产邻苯二酚菌株进行高通量筛选。结果:硫酸二乙酯的终浓度为0.1%,诱变时间为8 min的条件下,突变菌致死率为84.5%,突变效果最好。筛选培养基中吸光值(495nm)和富集培养基中菌液浊度值(OD630)的加和值较大的突变菌株产邻苯二酚能力高。通过诱变和筛选得到的菌株,产邻苯二酚浓度可达0.87mg/ml,比出发菌株的提高了262.5%。经过形态学和生理生化反应,初步鉴定该菌株属于假单胞菌属(Pseudomonas sp.)。结论:硫酸二乙酯诱变和96孔板筛选的方法能以高通量方式快速筛选出产邻苯二酚菌株。

Objective：Screen of strains producing catechol.Method：Strains producing catechol were screened using diethyl sulphate as mutagenizing reagent and 96 well plate screening method.Result：It showed that mutagenizing reagent content at ratio of 0.1%,at treatment time of 8min were suitable conditions for mutagenesis.Under specific conditions,the lethal rate was 84.5%.The mutant strains which showed high absorbance value in 495nm in sceening fermentation medium and high OD630 in enrichment medium would be better candidate for catechol-producing.Though mutagenizing and plate selection,the catechol yield of the strian was 0.87mg/ml,increasing by 262.5% compared to the original one.The strain was preliminary identified as Pseudomonas sp.by means of morphological,physiological and biochemical identification.Conclusion：Catechol producing strains could rapidly screened by high throughout screening using diethyl sulphate as mutagenizing reagent and 96 well plate screening method.