pAC-EPSILON
(Plasmid #53276)

• Purpose: Contains ctrE, crtB, and crtI genes of Erwinia herbicola (Pantoea agglomerans) Eho10, and the lcyE gene of Lactuca sativa and thereby produces epsilon-carotene in Escherichia coli
• Depositing Lab: Francis X Cunningham Jr
• Publication: Cunningham FX et al Plant J. 2005 Feb;41(3):478-92.

Backbone

• Vector backbone: pAC-LYC
• Backbone manufacturer: Francis X. Cunningham, Jr.
• Backbone size w/o insert (bp): 9476
• Total vector size (bp): 11729
• Modifications to backbone: A 2.28 kb PvuII-PvuII fragment with the Lactuca sativa (romaine lettuce) lycopene ɛ-cyclase was excised from the original cDNA library plasmid pDY4 [Cunningham FX Jr, Gantt E (2001) One ring or two? Determination of ring number in carotenoids by lycopene epsilon-cyclases. Proc Natl Acad Sci USA. 98, 2905-2910] and ligated in the Klenow-blunted SalI site of pAC-LYC. Orientation of the inserted lettuce lcyE cDNA was ascertained by restriction enzyme digestion.
• Vector type: low copy number bacterial cloning vector

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol, 25 μg/mL
• Growth Temperature: 30°C
• Growth Strain(s): Top10
• Growth instructions: Grow liquid cultures on a platform shaker at 28 degrees Celsius for 2-3 days in darkness for best epsilon-carotene production, or grow on agar plates at room temperature for 3-7 days.
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: lcyE
• Alt name: lycopene epsilon bicyclase
• Species: Lactuca sativa
• Insert Size (bp): 2249
• GenBank ID: AF321538.1
• Promoter: lac

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII (destroyed during cloning)
• 3′ cloning site: HindIII (destroyed during cloning)
• 5′ sequencing primer: none
• 3′ sequencing primer: none

Resource Information

• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

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

Depositor Comments

For better yield of low copy number pAC-based plasmids, grow liquid cultures on a platform shaker at ca. 30 degrees Celsius. When cultures reach early stationary phase, dilute 2-fold with growth medium, add spectinomycin (150 mg/liter), and "amplify" for several hours before harvest. For plasmid selection and maintenance in E. coli, use chloramphenicol at 30 mg/liter.