10XSTAT92E–luciferase
(Plasmid #37393)

• Depositing Lab: Norbert Perrimon
• Publication: Baeg et al Genes Dev. 2005 Aug 15;19(16):1861-70. Epub 2005 Jul 29.

Backbone

• Vector backbone: pUAST
• Backbone size w/o insert (bp): 8904
• Vector type: Insect Expression, Luciferase ; JAK/STAT reporter construct

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: SOCS36E enhancer fragment
• Alt name: 10X STAT92E binding sites
• Alt name: JAK/STAT reporter construct
• Alt name: SOCS36E
• Species: D. melanogaster (fly)
• Entrez Gene: Socs36E (a.k.a. Dmel_CG15154, CG15154, Dmel\CG15154, SOCS, SOCS36E, Socs, anon-EST:Liang-2.2, clone 2.2, dSOCS, dmSocs36E, socs36E, socs60E1)
• Promoter: Hsp70

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: multiple (see below for details) (unknown if destroyed)
• 3′ cloning site: multiple (see below for details) (unknown if destroyed)
• 5′ sequencing primer: pUAST-5 (5'-CTGCAACTAAAATCTGCCAAGAAG-3')

Resource Information

• Articles Citing this Plasmid:
  • 2 References

Terms and Licenses

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

Depositor Comments

Perrimon Lab plasmid #446

A 441-bp genomic fragment in the enhancer of SOCS36E containing two potential STAT92E-binding sites was amplified by PCR, using five different sets of oligos: (1) CTGCAGGAACCACTCAGAGTGCCTGCGTGT (PstI), GAATTCATACAAAACTGTCTTAGGTGTTTA (EcoRI); (2) CTGCAGGAACCACTCAGAGTGCCTGCGTGT (PstI), CTGCAGATACAAAACTGTCTTAGGTGTTTA (PstI); (3) GAATTCGAACCACTCAGAGTGCCTGCGTGT (EcoRI), GAATTCATACAAAACTGTCTTAGGTGTTTA (EcoRI); (4) AGATCTGAACCACTCAGAGTGCCTGCGTGT (BglII), AGATCTATACAAAACTGTCTTAGGTGTTTA (BglII); (5) GCGGCCGCGAACCACTCAGAGTGCCTGCGTGT (NotI), GCGGCCGCATACAAAACTGTCTTAGGTGTTTA (NotI).

Each amplified genomic fragment containing different restriction enzyme sites was sequentially subcloned into pUAST. The genomic fragment amplified using the first set of oligos was subcloned into the PstI/EcoRI sites of pUAST to generate 2XSTAT92E. The genomic fragment amplified using the second set of oligos was subcloned into the PstI site of 2XSTAT92E to generate 4XSTAT92E. The genomic fragment amplified using the third set of oligos was subcloned into the EcoRI site of 4XSTAT92E to generate 6XSTAT92E. The genomic fragment amplified using the fourth set of oligos was subcloned into the BglII site of 6XSTAT92E to generate 8XSTAT92E.

Next, the hsp70 minimal promoter element was amplified from pUAST by PCR using oligos GCGGCCGCAGCGGAGACTCTAGCGAGCG (NotI) and CTCGAGAATTCCCTATTCAGAGTTCT (XhoI). This
hsp70 minimal promoter was subcloned into the NotI/XhoI sites of 8XSTAT92E to generate 8XSTAT92E–hsp70. Again, the genomic fragment amplified using the fifth set of oligos was subcloned into the NotI site of 8XSTAT92E–hsp70 vector to generate 10XSTAT92E–hsp70.

Finally, an XhoI/XbaI fragment containing the firefly luciferase gene from the pGL3–luciferase vector (Promega) was subcloned into the XhoI/XbaI sites of 10XSTAT92E–hsp70 to generate 10XSTAT92E–luciferase.