CC1215
(Bacterial strain #79062)

• Purpose: BL21(DE3)-based E. coli purine auxotrophic host strain used for the preparation of PurE proteins in a purE background.
• Depositing Lab: T. J. Kappock
• Publication: Kappock unpublished strain (unpublished)

Backbone

• Vector backbone: none

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): CC1215 (purE)
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: none
• Promoter: none

Cloning Information

• Cloning method: Unknown
• 5′ sequencing primer: ODN 1161: 5'-CGGATGACGTTGAGCTGATTCATTTTCC
• 3′ sequencing primer: ODN 1164: 5'-CAGCTCACCTAAACGATCAAACAC

Resource Information

• Supplemental Documents:
  • Primer information for strain verification
  • PCR strain verification for CC1215

Terms and Licenses

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

Depositor Comments

Genotype = pur
Precursor strain = BL21(DE3)

This strain was built by Kelly L. Sullivan in the lab of T. J. Kappock. It is used to produce PurE proteins, including low-activity mutants, in a zero-background host lacking a functional purE1 gene. Both N5-CAIR mutase (purE1; EC 5.4.99.18) and AIR carboxylase (purE2; EC 4.1.1.21) mutants have been successfully overproduced in this strain.

Purine auxotrophy results from a defect in the de novo biosynthesis of purine nucleotides. The auxotrophy can be functionally complemented with adenine, hypoxanthine, or other substrate for the purine salvage pathway. There is no evident requirement for His or thiamine.

Strains lacking a functional purE1 gene excrete AIR nucleoside (AIRs) which spontaneously forms a brown polymer. Culture media and plates may have a darker pigmentation unless a functional purE (either purE1 or purE2) is present or a purine base, which represses production of de novo biosynthesis enzymes, is supplied.

The deletion cassette ΔpurE736::kan from Keio strain JW0512 was transduced (using P1) to disrupt the target gene in BL21(DE3). Subsequent attempts to excise the cassette were unsuccessful.