Optimizing Enzyme Engineering for Single Cell Genomics

Overview

Single cell genomics is a powerful approach that offers comprehensive insights into biology by probing for genomic information from individual cells. Due to the limited genetic materials in a single cell, it remains challenging to implement new single cell genomic profiling approaches. Novel enzymes for amplifying and cloning DNA with enhanced sensitivity and specificity could address these challenges. In this study, we establish workflows for engineering enzymes for single cell genomics application. we used T4 DNA Ligase and Pfu DNA Polymerase as models to determine optimal methods and conditions to express and purify these enzymes. Various concentrations of inducers (IPTG), induction times, and growth temperatures of the bacterial strain carrying the gene for these enzymes were tested to optimize enzyme production and quality. The proteins were purified, and enzyme yield and activity were measured for each growth condition. Results indicate that a 10 µM concentration of IPTG gave the best yield for both enzymes, and shifting the cultures to lower temperature after IPTG induction yielded better enzyme quality due to increased solubility.

Competition history

• AJAS 2020 Category not listed

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