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微流控芯片监测绿色荧光蛋白在枯草芽孢杆菌中的表达

来源:中科院微生物研究所 2009-09-01 18:25

生物工程学报 25 July 2009, 25(7):1077~1081

微流控芯片监测绿色荧光蛋白在枯草芽孢杆菌中的表达

董惠钧1*, 付景林2*, 李永泉3, 姜俊云1

1 鲁南制药集团股份有限公司研发中心, 临沂276005

2 浙江大学微分析系统研究所, 杭州310058

3 浙江大学生命科学学院, 杭州310058

摘 要: 目前主要使用激光共聚焦扫描显微镜观察绿色荧光蛋白的表达, 但需要昂贵的仪器并耗费大量时间。本研究开发了一种新型激光诱导的微流芯片检测系统来监测绿色荧光蛋白在枯草芽孢杆菌中的表达。该系统主要由激光装置、光路系统、微流控芯片、光电倍增管和计算机处理系统等5部分组成。对该系统的测试结果显示, 随着诱导强度的增强监测信号峰也随之增强, 并且与激光共聚焦显微镜观察的结果一致。利用该芯片系统能够快速准确地筛选和鉴定用绿色荧光蛋白作为标记的细胞克隆, 可以替代PCR鉴定方法。但该系统仅仅能够监测表达强度, 不能够满足蛋白定位等高水平研究, 因此, 该系统适合应用于环境的微生物监测、药物筛选和其他无需观察蛋白定位等研究。 关键词: 微流控芯片, 绿色荧光蛋白, 激光诱导荧光, 枯草芽孢杆菌

Microfluidic chip for detecting the expression of green fluorescent protein in Bacillus subtilis

Huijun Dong 1*, Jinglin Fu 2*, Yongquan Li 3, and Junyun Jiang1

1 Research and Development Center, Lu’nan Pharmaceutical Group Inc. Ltd., Linyi 276005, China

2 Institute of Microanalytical Systems, Zhejiang University, Hangzhou 310058, China

3 College of Life Sciences, Zhejiang University, Hangzhou 310058, China

Abstract: Laser scanning confocal microscope (LSCM) is currently the only equipment to observe fluorescence. However, this technique has disadvantages such as high cost and long test process. In this study, we developed a new system of laser-induced fluorescence (LIF) for microfluidic chip applied to detecting the expression of green fluorescent protein (GFP) in Bacillus subtilis. This novel system was comprised of laser device, optics unit, microfluidic chip, photomultiplier and computer treatment unit. The tests indicated that microfluidic chip could detect the expression of GFP as sensitively as LSCM in Bacillus subtilis. Moreover, this LIF detection system could instead of PCR to identify the positive clone in this special case. Nevertheless, the LIF system only was suitable to detect the fluorescent strength of GFP, and could not meet the request of some cases for example protein location. Therefore, this system will be applied in environmental detection with microbe, drug discovery and other cases.

Keywords: microfluidic chip, green fluorescent protein (GFP), laser-indced fluorescence (LIF), Bacillus subtilis

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