Expressing proteins in bacteria is critical to studying protein functions. However, protein expression and purification is a multi-day process. The NEB PURExpress^® Cell Free E. coli protein synthesis system claims to enable protein expression in only 2-4 hours. Another appealing aspect of this kit is that it allows for protein expression using PCR products as templates for in vitro protein expression. This eliminates the need for laborious biologic cloning of the cDNA into a plasmid vector. To expedite protein function analyses, our laboratory used the NEB PURExpress® system to express C. elegans proteins. Preliminary experiments demonstrated that three C. elegans proteins that were expressed using this kit all failed to exhibit a single band corresponding to their expected sizes upon western blotting. Trials conducted under different experimental conditions were unable to yield proper protein expression using this kit. On the other hand, the conventional method of bacterial protein expression in BL21 E. coli cells were more fruitful in yielding proper protein expression of two out of the three above-mentioned C. elegans proteins. Notably, two predominant bands were produced using the conventional method with their bands corresponding to their expected protein sizes. One C. elegans protein was difficult to express in bacteria, likely due to its known low expression level and its toxicity to bacteria. Overall, our data indicate that the conventional method of protein expression yielded more favorable results than using the NEB PURExpress® system while expressing C. elegans proteins. Therefore, we conclude that the conventional method of protein expression in bacteria is likely preferrable to using the more expensive NEB PURExpress® system while expressing C. elegans proteins.

Gregory Palomar

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