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Analytica Chimica Acta
Volume 1040, 2018, Pages 105-111

A fast and visual method for duplex shrimp pathogens detection with high specificity using rapid PCR and molecular beacon

Cheng Qiana, Rui Wanga, Cui Wua, Liu Wanga, Zunzhong Yea, Jian Wua, Feng Jib

College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou, 310058, China.

Abstract

Molecular diagnosis of genome is one of the major methods for pathogens detection. The commonly used PCR method can realize an exponential amplification of the target gene but is time-consuming. In this work, we proposed a duplex and visual method using rapid PCR combined with molecular beacons to specifically detect two kinds of shrimp pathogens in one reaction tube. We only need to observe the fluorescence change of the reaction tube with naked eye to determine the result. A home-made automatic transfer equipment allows reaction tubes shuttling quickly between two water baths to achieve rapid PCR amplification. A simple device was also designed to present the detection results easily determined with naked eye. This duplex and visual detection method is fast, low-cost and of high specificity. From DNA extraction to results judgment, only 15 min was enough. Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) and Vibrio parahaemolyticus (VP) are two common shrimp pathogens which were chosen as our detection objects. This method may give a possibility to conduct end-point visual duplex detection, which may make a positive influence on the pathogen prevention.

Graphical abstract

Keywords: Duplex and visual detection, Rapid PCR, Molecular beacon, Shrimp pathogens.

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