Imidacloprid is a systemic insecticide used to control termites, fleas, and sucking insects by disrupting their central nervous system. This chemical is found in many products and is beneficial in people’s homes, pets, and crops. However, an increased amount of exposure to imidacloprid is toxic to humans and detrimental to the environment and wildlife. According to the National Pesticide Information Center, brief exposure to imidacloprid may cause eye irritation, confusion, or vomiting. Additionally, this insecticide is traced in groundwater due to the drainage from soil which subsequently terminates fish and other sea animals. Thus, it is important to continuously detect levels of imidacloprid in soil and water to manage this public health issue. The goal of our lab is to create DNA-based biosensors for imidacloprid using the method called Systematic Evolution of Ligands by Exponential Enrichment, or SELEX. This process identifies specific DNA aptamers from a random DNA library that will bind to the target of interest (imidacloprid), with high affinity and high specificity. In every round, we introduced the imidacloprid target to collect all the specific DNA that binds to it. We lowered the concentration of the target every few rounds in order to isolate high binding affinity aptamers. We have completed 25 rounds of SELEX and the target concentrations were decreased from 100 uM to 25 nm. The concentration of the single-stranded DNA was measured and 100 pmol was used as the starting library for the next SELEX round. Finally, we incorporated the purified PCR product (750 ng/uL) directly into the plasmid vector. The plasmids were then transformed into competent cells, and positive colonies containing recombinant plasmid DNA were screened via blue/white screening. We randomly picked 22 colonies and cultured them in LB media. We isolated plasmids from the cultured cells and sent the samples to the DNA sequencing service facility. We received 21 sequences back and are currently testing the biosensors in fluorescence assay. These findings will help us to design and develop a DNA aptamer-based biosensor for the detection of imidacloprid.

Development of DNA-based fluorescent biosensor for detection of pesticide Imidacloprid

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.