Diagnosis of Sugarcane White Leaf Disease Using the Highly Sensitive DNA Based Voltammetric Electrochemical Determination

Abstract

A simple and highly sensitive analysis by electrochemical voltammetry has been developed for diagnosis of the most destructive crop disease in Thailand known as sugarcane white leaf (SCWL). Determination of the corresponding DNA interaction has been obtained from the voltammetric signals of electroactive redox methylene blue (MB) by means of cyclic and differential pulse voltammetry. In this study, a chitosan-modified glassy carbon electrode (GCE) was created by self-assembly to produce electrostatic platform for effective immobilization of the DNA. Fabrication of SCWL-DNA hybridization detection system was performed by immobilizing the ssDNA probe as a specific sensor onto chitosan-modified GCE. Hybridization of complementary DNA from the real samples could then be detected by its respective MB signal. This fabricated DNA probe sensor was shown to be capable for discriminative identification among the DNAs from SCWL plants, mosaic virus infected sugarcane and healthy sugarcane plants. Relationship between the specific hybridization signal and DNA target concentration was also observed under optimal condition. The detection limit of 4.709 ng/µl with the regression coefficient (R 2 ) of 0.998 and overall RSD of 2.44% were obtained by response curve fit analysis. The actual SCWL-ssDNA immobilization and hybridizing event were subsequently confirmed by an observation under atomic force microscope. Thus these experiments demonstrate the first successful and effective DNA based voltammetric electrochemical determination for a verification of the specific pathogenic infection within plants from the real epidemic field.

The Incorporation of Amplified Metal-Enhanced Fluorescence in a CMOS-Based Biosensor Increased the Detection Sensitivity of a DNA Marker of the Pathogenic Fungus Colletotrichum gloeosporioides

“…The light signal was higher by 18.75-, 11.11-, 5.5-, 11.25-, and 3.75-times in the improved protocol for all the tested concentrations of the target DNA strand of 1000, 100, 10, 3.3, and 2 nM, respectively. The developed CMOS-based biosensor system demonstrated comparable detection sensitivity with other optical-based [ 73 , 74 ], electrochemical-based [ 75 , 76 ], and mass-balance-based [ 77 , 78 ] biosensor platforms for pathogen detection in plants. The PCR technology demonstrates a more sensitive DNA detection; however, it is not suitable for real-time monitoring on agricultural sites.…”

Nanostructural Characterization of Glutathione-S-Transferase Immobilizing Chitosan Modified Screen Printed Carbon Electrode by Atomic Force Microscopy

“…As it is soluble in weak acids and forms quite stable hydrogel film which make it suitable matrix for various micro-fabrications. Our previous study on the modification of a screen printed carbon electrode by surface incorporation with chitosan has demonstrated one among potent application for obtaining an effective and affordable electrochemical implement [1]. With reference to such excellent advantages, it was further used to achieve another successful immobilization of enzyme in this present work to gain an effective specific biosensor.…”

“…Recently, researchers based on electrochemical biosensor have been attractively considered as potential successors to the development of new methodologies for economical and real-time monitoring of environmental pollutants and also for prevention of toxic materials in the environment [1] The main functional activity belongs to the oxidising and reducing ability of the selective bio-recognition elements that make a high throughput screening of analytical target possible in a reliable manner within a fraction of seconds. Therefore a critical step for the development of biosensor is the selection of a proper bio-recognition element and the immobilization of this element to the surface of a working platform.…”

“…The fact that nanotopographical surfaces influence charge density and electric field strength, quantitative characterization of surface morphology is thus very important. To date, AFM has been employed progressively for exploring the surface morphological features especially in the research area of surface engineering [1,9].…”

Development of Specific Electrochemical Biosensor Based on Chitosan Modified Screen Printed Carbon Electrode for the Monitoring of Captan Fungicide

“…Typical peak curve of the reductive and reverse oxidative current vs. electrode potential indicating ideal reversible reaction governed by freely diffusion of Fe +3 /Fe +2 to the planar surface was gained in each electrode at about 0.09 to 0.21 V with the peak separation around 0.1 V at 100 mV s -1 scan rate and a potential window of −0.6 to 0.6 V as shown in Fig.3. The oxidation/reduction peak currents of SPCE-Chi and SPCE-Chi-GST were almost 2-3 times bigger than that of original bare SPCE (curve a) as a successive superb improvement in electroactive response due to the surface furnishing by an enriched cationic chitosan biopolymer [8,9,10]. A high rate of electron transfer process was thus accelerated via this highly conductive chitosan composing as demonstrated by the curve b for SPCE-Chi and curve c for SPCE-Chi-GST.…”