CRISPR Interference Knockdown of Ampicillin Resistance Gene in Escherichia coli

Overview

Objectives/Goals Synthesis of a CRISPRi plasmid to silence the ampicillin resistance gene in Escherichia Coli. Methods/Materials Constructed a gRNA scaffold inside the CRISPRi plasmid encoding for a 20bp sequence of the ampicillin resistance gene, transformed E. coli with the plasmid through heat-shock, plated bacteria onto chloramphenicol LB agar plates (plasmid contained a chloramphenicol resistance gene), swabbed bacteria onto new LB agar plates and placed one ampicillin disc onto each plate, measured the zone of inhibition around each disc after a 24-hour growth period in incubator. Results The measurements of the zones of inhibition surrounding the ampicillin disc on the plate containing the transformed E. coli showed susceptibility to the antibiotic while there was no zone of inhibition on the E. coli not transformed with the CRISPRi plasmid which demonstrated a resistance to the antibiotic. Conclusions/Discussion The constructed CRISPRi plasmid showed efficacy to the suppression of the ampicillin resistance gene in Escherichia Coli. This demonstrated then value of utilizing CRISPR or CRISPRi in the plasmid form to modify/regulate gene sequences and expressions in organisms, including the repression of antimicrobial resistance genes in bacteria which can help treat infectious diseases with more efficiency.

Awards (1)

• Honorable Mention

Competition history

• CSEF 2018 Microbiology (General) · Entry S1613

Resources

• CSEF project summary ↗

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