生物工程學報 25 July 2009, 25(7):1077～1081

微流控芯片監(jiān)測綠色熒光蛋白在枯草芽孢桿菌中的表達

董惠鈞1*, 付景林2*, 李永泉3, 姜俊云1

1 魯南制藥集團股份有限公司研發(fā)中心, 臨沂276005

2 浙江大學微分析系統(tǒng)研究所, 杭州310058

3 浙江大學生命科學學院, 杭州310058

摘 要: 目前主要使用激光共聚焦掃描顯微鏡觀察綠色熒光蛋白的表達, 但需要昂貴的儀器并耗費大量時間,。本研究開發(fā)了一種新型激光誘導的微流芯片檢測系統(tǒng)來監(jiān)測綠色熒光蛋白在枯草芽孢桿菌中的表達。該系統(tǒng)主要由激光裝置,、光路系統(tǒng),、微流控芯片、光電倍增管和計算機處理系統(tǒng)等5部分組成,。對該系統(tǒng)的測試結(jié)果顯示, 隨著誘導強度的增強監(jiān)測信號峰也隨之增強, 并且與激光共聚焦顯微鏡觀察的結(jié)果一致,。利用該芯片系統(tǒng)能夠快速準確地篩選和鑒定用綠色熒光蛋白作為標記的細胞克隆, 可以替代PCR鑒定方法。但該系統(tǒng)僅僅能夠監(jiān)測表達強度, 不能夠滿足蛋白定位等高水平研究, 因此, 該系統(tǒng)適合應用于環(huán)境的微生物監(jiān)測,、藥物篩選和其他無需觀察蛋白定位等研究,。 關鍵詞: 微流控芯片, 綠色熒光蛋白, 激光誘導熒光, 枯草芽孢桿菌

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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