Automated, high-throughput DNA extraction protocol for disposable label free, microfluidics integrating DNA biosensor for oil palm pathogen, Ganoderma boninense. (May 2020) - Ayoib, Adilah
Graphical abstract: Highlights: Basal stem rot disease caused by pathogenic fungus Ganoderma boninense in the oil palm. Automated DNA extraction on microfluidics for quick, disposable, label-free detection. G.boninense & other pathogenic fungus isolated from mushroom & DNA fractionated. Successful fabrication with <0.4 mm variance in dimension in the design lithography. Comparable result with synthetic ssDNA of G. boninense & contrast with other fungus. Abstract: Basal Stem Rot and Upper Stem Rot diseases caused by pathogenic fungus Ganoderma boninense continue to be a major plight in the palm oil industry. Despite continuous research in combating the problem, resolution remains stagnant. Here, developed an automated, high-throughput DNA extraction protocol on microfluidics device for a quick, disposable, label-free detection, within 2 h of assessment. Microfluidics was designed using AutoCAD software, fabricated on microscopic glass substrate using negative photoresist (SU-8 2015) and molded with a biopolymer silicone, Polydimethylsiloxane. G. boninense and unknown pathogenic fungus isolated from rotten mushroom were grown and fractions of extracted DNA were pooled and analyzed for comparison along with synthetic ssDNA of G. boninense . Results from LPM and HPM show successful fabrication with ≤0.1 mm variance between the dimensions in the design before and after lithography process. The PDMS microfluidics show no leakage when run with DNA samples. Analyses from I-V measurement, UV–vis, FTIR, and PCR show comparable results between extracted and synthetic ssDNA of G. boninense and a contrast with the unknown pathogenic fungus, indicating a successful DNA extraction protocol via microfluidics for label-free identification of G. boninense. Optimization of DNA extraction can be further devised for applicability on lab-on-a-chip devices.