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Electrochemical detection of β-1,3-glucanase gene from transgenic capsicums using asymmetric PCR generated by a detecting probe and an anchoring probe

Wu, Guofan, Wang, Zhihua, Zhang, Huini, Yang, Ning, Du, Jie, Lu, Xiaoquan, Kang, Jingwan
Journal of biotechnology 2010 v.145 no.1 pp. 1-8
Capsicum, transgenic plants, beta-glucanase, transgenes, polymerase chain reaction, DNA probes, electrochemistry, electrodes, recombinant DNA, genetic vectors
A design for recognition of β-1,3-glucanase gene (Glu) specific sequence based on probe extension was described. The detecting probe DNA and the anchoring prober were hybridized with the same target DNA firstly, then the probes were extended by DNA polymerase reaction. After that the double strand DNA was denatured, and the extended detecting probe was immobilized on a glassy carbon electrode via nanoparticle gold (AuNP). In electrochemical detection (cyclic voltammetry, CV and differential pulse voltammetry, DPV), an increased peak current (i p) of the indicator (methylene blue, MB) was obtained compared with the probe without extension. Three differently long DNAs of Glu specific sequence were employed as the target: oligonucleotide acid, molecular cloning vector DNA and total genome DNA of transgenic capsicum. The estimated DPV detection limits for three targets of oligonucleotide, the molecular cloning vector DNA and genome DNA were 2.6×10⁻¹³, 6.0×10⁻¹³ and 8.0×10⁻¹³ mol l⁻¹ respectively.