基于荧光定量PCR技术构建水源地典型致嗅物质2-甲基异莰醇的评估方法及其应用  

Evaluation of typical odorant 2-methylisoborneol based on real time qPCR in source water and its application

作　　者：徐晓庆 苏命[2] 朱宜平 崔长征[1] MUHAMMAD Suruzzaman 徐亚楠 于建伟[2] 杨敏[2] XU Xiaoqing;SU Ming;ZHU Yiping;CUI Changzheng;MUHAMMAD Suruzzaman;XU Yanan;YU Jianwei;YANG Min(State Environmental Protection Key Laboratory of Environmental Risk Assessment and Control on Chemical Process,School of Resources and Environmental Engineering,East China University of Science and Technology,Shanghai 200237,China;State Key Laboratory of Environmental Aquatic Chemistry,Research Center for Eco-Environmental Science,Chinese Academy of Sciences,Beijing 100085,China;Shanghai Chengtou Raw Water Co.Ltd.,Shanghai 200125,China)

机构地区：[1]华东理工大学资源与环境工程学院,国家环境保护化工过程环境风险评价与控制重点实验室,上海200237 [2]中国科学院生态环境研究中心,环境水质学国家重点实验室,北京100085 [3]上海城投原水有限公司,上海200125

出　　处：《环境工程学报》2020年第11期3208-3215,共8页Chinese Journal of Environmental Engineering

基　　金：国家水体污染控制与治理科技重大专项(2017ZX07108-002-02);国家自然科学基金资助项目(51878649)。

摘　　要：我国地表水源中由丝状蓝藻代谢产生2-甲基异莰醇(2-MIB)导致的嗅味问题十分普遍。由于传统显微镜检测方法无法鉴别产嗅藻种,不能满足水源水质管理要求,故有必要构建能特异性表征水体产嗅潜力的检测方法。由于丝状藻代谢产生2-MIB主要受其合成功能基因调控,设计了2-MIB功能基因引物,经引物特异性检验及PCR条件优化后,构建了基于2-MIB功能基因的定量PCR方法,并采用实际环境样品进行测试与验证。结果表明:引物特异性良好,定量PCR方法能有效检测2-MIB功能基因,并绘制了标准曲线,得出检测限为8.44×102 copies·L-1;实际样品测试结果显示其2-MIB功能基因浓度在2.09×107~1.94×1010 copies·L-1范围内,与基于仪器分析方法测定的2-MIB浓度符合线性关系(R2=0.63,P<0.01),表明定量方法可行。该方法具有灵敏性高、特异性好的特点,能特异性检测产嗅基因,可应用于水源地中产嗅潜力评估与预警。Two-methylisoborneol(2-MIB) is a typical secondary metabolite released from cyanobacteria which causes off-flavor odor problem in surface water across China. Microscopy is the traditional method to identify such cyanobacteria. However, merely it is troublesome to distinguish between 2-MIB producers and nonproducers by morphological structure and cannot meet the requirement of the source water quality management.It is necessary to build the test method which can specifically characterize the odor-producing capacity of waterbody. Due to the functional gene regulation for 2-MIB yield from cyanobacteria, the primers of 2-MIB function gene were designed and its specificity test was performed, then the PCR conditions were optimized.Real-time quantitative polymerase chain reaction(qPCR) can be an alternative method to evaluate 2-MIB syntheses which is controlled by 2-MIB functional gene. Here, 2-MIB identification primers(MIBF/MIBR)have been designed and tested with field samples. A good standard curve was established, R2=0.999 5, P<0.01,and the detection limit was 8.44×102 copies·L-1. The concentrations of 2-MIB gene of field samples were between 2.09×107 copies·L-1 to 1.94×1010 copies·L-1, which were significantly conformity with 2-MIB concentrations measured by GC-MS(R2=0.63, P<0.01). The high sensitivity and specificity of this qPCR-based method suggests that it can effectively evaluate the risk of 2-MIB occurrence and able to monitor source water quality management.

关 键 词：定量PCR 2-甲基异莰醇 产嗅功能基因 丝状蓝藻 饮用水 

分 类 号：X832[环境科学与工程—环境工程]