Q. What proteins and nucleic acids can be produced in Clean Genome® E. coli?
A. Clean Genome® E. coli has so been used to produce ~20 diverse proteins plus single-chain antibodies, retroviral, lentiviral and shRNA plasmids, and a DNA vaccine.
Q. How do I achieve maximum expression of my target protein?
A. Clone your target gene into the pSX2 expression vector and express it in one of the Clean Genome® E. coli strains.
Q. What concentration of IPTG should I use for inducing my protein or nucleic acid expression?
A. The optimum concentration of IPTG may vary with the target product. We recommend testing IPTG concentrations from 5 µM to 500 µM for each target product.
Q. How do I efficiently produce plasmids that contain secondary structure such as viral Long Terminal Repeats (LTR) or short-hairpin RNA (shRNA) etc?
A. Use the Clean Genome® E. coli ΔrecA or Clean Genome® E. coli LowMut ΔrecA strain for production. These strains stabilize such challenging plasmids as pT-ITR-IL2, an adeno-associated virus vector that contains particularly extensive secondary structure.
Q. Why do I care about E. coli insertion elements?
A. Insertion (IS) elements are present in all E. coli strains except for Clean Genome® E. coli. IS elements can transpose into any DNA present in the cell, including the genome and the plasmid carrying the gene and regulatory elements for the product you want to make. Transposition of IS elements into the promoter can reduce or eliminate expression of the product, and transposition into the product gene could mutate it, generating defective product.
Q. How do I determine whether my DNA of interest contains IS elements?
A. Use the Insertion Sequence Detection Kit. This kit is designed to detect IS contamination in DNA you make as well as in commercially produced DNAs, where it is typically found.
Q. How can I produce nucleic acids that are free of Insertion Elements?
A. Produce your nucleic acids in Clean Genome® E. coli.
Restriction, Modification and Methylation
Q. Do the Clean Genome® E. coli strains express the Dam or Dcm methylases?
A. Yes, they are dam+ and dcm+.
Q. What host restriction systems have been deleted from the Clean Genome® E. coli strains?
A. They are ΔhsdR, ΔhsdM and ΔhsdS.
Q. What host modification systems have been deleted from the Clean Genome® E. coli strains?
A. They are ΔmcrA, ΔmcrB, ΔmcrC, and Δmrr.
Acquiring Clean Genome®
Q. How do I purchase Clean Genome® E. coli strains?
A. Clean Genome® E. coli strains are available for a fee for licensing, but they are not available for purchase.
Q. What if I just want to use Clean Genome® E. coli for academic research?
A. Congratulations, you made an excellent choice. Prior to shipping, we will need to receive a Limited Use License Agreement signed by an individual from your institution who is authorized to enter into such agreements.
Q. What if I want to use Clean Genome® E. coli for commercial purposes?
A. Congratulations, you made an excellent choice. A commercial license is required for commercial use, please contact firstname.lastname@example.org to discuss.
Q. Is there a way to obtain a strain of Clean Genome® E. coli that has a genotype not available in your current product list?
A. We have an extensive library of Clean Genome® E. coli strains with various additional genetic modifications, please contact email@example.com for your specific needs.