pJH299
(Plasmid #13325)

• Purpose: recombination substrate
• Depositing Lab: Martin Gellert
• Publication: Hesse et al Genes Dev. 1989 Jul . 3(7):1053-61.

Backbone

• Vector backbone: pUC13

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: recombination substrate

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: SalI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: n/a

Terms and Licenses

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

Depositor Comments

pJH299 is derived from pJH298. pJH298 is described in Lieber et al., 1988. Briefly, the plasmid backbone is a fusion of pUC13 and the early region of the polyoma virus genome, which allows the plasmids to replicate autonomously in E. coli and murine cells, respectively. Various DNA fragments pertinent to this recombination assay are inserted into the multipurpose cloning site of pUC13, including a prokaryotic promoter, the structural gene encoding chloramphenicol acetyltransferase (cat), and the intervening transcription terminator. The terminator is flanked on the promoter-proximal side by a unique SalI site and on the cat gene side by a unique BamHI site. DNA fragments containing V(D)J recombination signals are inserted into these unique sites. (These signals are the only DNA segments related to Ig genes or TCR genes included on the plasmid). A 39-bp fragment containing a 12-signal, created by annealing of synthesized complementary oligonucleotides with protruding ends compatible with SalI, is inserted into the SalI site of the backbone. A 49bp fragment containing a 23-signal, made similarly but with protruding ends compatible with BamHI, is inserted into the BamHI site of the backbone.  pJH298 contains signals bearing the consensus versions of heptamers and nonamers.

pJH299 contains signals bearing the consensus versions of heptamers and nonamers.