RF12
(Bacterial strain #62077)

• Purpose: BL21(DE3)-based E. coli amino acid auxotrophic host strain used for selective isotope labeling. trpA trpB genes deleted.
• Depositing Labs: Robert Gennis, Toshio Iwasaki
• Publication: Lin et al Methods Enzymol. 2015;565:45-66. doi: 10.1016/bs.mie.2015.05.012. Epub 2015 Jun 10.

Backbone

• Vector backbone: none

Growth in Bacteria

• Bacterial Resistance(s): None
• Growth Temperature: 37°C
• Growth Strain(s): RF12 (trpA trpB )
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: none

Resource Information

• Supplemental Documents:
  • PCR primers
  • PCR data RF12

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Genotype = trpA trpB
Precursor strain = BL21 CodonPlus (DE3)-RIL
Selective amino acid labeling (and/or requirement) = Trp##

##RF12 strain requires Trp for growth in M63 minimal medium. For effective Trp labeling of a recombinant protein, the presence of 0.4-1 mM Tyr in growth medium to repress the tyrB gene is recommended. This strategy can only be applicable for a short-term cultivation but not suitable for a long-term cultivation for heterologous expression of foreign genes [Methods 55, 370-378 (2011)].

Each target gene was deleted from the chromosome of BL21 (DE3) E. coli strain using λ-Red recombination system.

Supplemental documents contain a list of PCR primers used for verification of each knocked-out gene as well as an image showing PCR results for this strain.

This work is supported in part by JSPS-NSF International Collaborations in Chemistry (ICC) research grant.