CJ22 (miR-1003 perfect site)
(Plasmid #17762)

• Depositing Lab: David Bartel
• Publication: Ruby et al Nature. 2007 Jul 5. 448(7149):83-6.

Backbone

• Vector backbone: pMT-puro
• Backbone manufacturer: David Sabatini Lab
• Vector type: Insect Expression, Luciferase ; Drosophila metallothionein gene promoter drives expression
• Selectable markers: Puromycin

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: miR-1003 perfect site
• Species: D. melanogaster (fly)
• Tag / Fusion Protein:
  • renilla luciferase (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ sequencing primer: MT forward

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Renilla Luciferase Limited Use Label License
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Luciferase-reporter inserts were made by annealing oligonucleotides with their reverse complements, leaving overhangs for the indicated restriction sites (lower case): miR-1003-ps (gagctcCTGTGAATATGTAAATGTGAGAactagt) Annealed oligos were ligated into SacI/SpeI-cleaved pIS2. These plasmids were linearized with HindIII, polished with Klenow enzyme to create blunt ends, and digested with NotI to excise the Renilla luciferase gene with the modified UTR from the remainder of pIS2. The gel-purified Renilla gene fragment was then ligated into pMT-puro between EcoRV and NotI sites for copper-induced expression in S2 cells.