Removal Of Contaminating Nucleic Acids From Recombinant Protein Preparations

April 18, 2018

University of Birmingham
Biosciences
Research Assistant

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Company:

MilliporeSigma

Product Name:

Benzonase Nuclease

Catalog Number:

70746-3

I have used Merckmillipore Benzonase nuclease to remove DNA and RNA from recombinant protein expressed in E. coli BL21 cells. Treatment with benzonase nuclease treatment with cell lysis protocol greatly reduced viscosity and improved flow rates of protein preps through chromatography columns. Got much pure proteins.

Experimental Design and Results Summary

Application

Removal of nucleic acids from recombinant proteins during protein extraction and purificataion

Starting Material

1 gram of E.coli BL21 cell pellets expressing protein treated with Novagen BugBuster reagent

Protocol Overview

Protocol is very simple. I regularly use Novagen BugBuster cell lysis protocol for extraction of proteins. Follow exactly the Novagen BugBuster lysis protocol. Once you add BugBuster reagent to 1X concentration to your cell pellets, then add 30U/ml Benzonase nuclease and incubate the mix for 1 hr at room temperature. Then follow next steps of protein purification as mentioned in the Novagen BugBuster protocol.

Tips

Avoid frequent freeze and thaw cycles of Benzonase nuclease. Make sure that monovalent cations in the buffer/lysis reagents should be less than 50 mM

Results Summary

Satisfactory results from Benzonase nuclease treatment. I was experiencing viscous lysate and flow through chromatography column for protein purification was taking much time with less yield. Treatment with Benzonase nuclease reduced viscosity and removed almost all nucleic acid contaminants from my protein preps. Quality was checked by Nanodrop spectrometer for Absorbance at 280 nm.

DOI or PMID #

N/A

Additional Notes

None

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Summary

The Good

Simple protocol and can be adapted any type of protein extraction and purification protocols

The Bad

Slightly expensive enzyme to buy for small scale protein purification

The Bottom Line

Overall, very much satisfied with using benzonase nuclease during my protein purification especially problem of reduced flow through in the chromatography columns was solved.

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