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pNiFty3 - mIFN-β promoter - AP-1 - ZeocinR - SEAP
pNiFty3-A-SEAP plasmid is composed of three key elements: the mouse interferon beta minimal promoter, five AP-1 transcription factor binding sites and a SEAP (Secreted alkaline phosphatase) reporter gene.
pNiFty3-A-SEAP plasmid is selectable with Zeocin™ in both E. coli and mammalian cells, and can be used to generate stable clones.Back to the top
Transcription factor binding sites: AP-1 (5x)
Minimal Promoter: mouse IFNβ promoter
Reporter Gene: SEAP
pNiFty3-A-SEAP plasmid is provided as 20 µg lyophilized DNA with 4 pouches of Fast-Media® (2 TB and 2 Agar), containing Zeocin™.
Products are shipped at room temperature and should be stored at -20°C.Back to the top
The proximal promoters are shorter than 500 bp and contain transcription factor binding sites. Upon stimulation in 293 cells, their expression level remains undetectable. With the addition of repeated TFBS, the proximal promoters become inducible by the appropriate stimulus and drive the expression of the reporter gene.
IFN-β promoter: the mouse IFN-β minimal promoter comprises several positive regulatory domains that bind different cooperating transcription factors such as NF-kB, IRF3 and IRF7 .
Transcription factor binding sites (TFBS)
AP-1 binding site: Activator protein 1 (AP-1) is a transcription factor activated by most PRRs. AP-1 is a heterodimeric complex composed of members of Fos, Jun and, ATF protein families. AP-1 binds to the TPA responsive element (TRE: ; TGAG/CTCA) . AP-1 activation in TLR signaling is mostly mediated by MAP kinases such as c-Jun N-terminal kinase (JNK), p38 and extracellular signal regulated kinase (ERK).
SEAP reporter gene: Secreted alkaline phosphatase (SEAP) is a reporter widely used to study promoter activity or gene expression. SEAP expression can be rapidly and readily measured in supernatants of transfected cells. SEAP levels can be evaluated qualitatively with the naked eye and quantitatively using SEAP detection media, such as HEK-Blue™ Detection system or the SEAP Reporter Assay.
1. Vodjdani G. et al., 1988. Structure and characterization of a murine chromosomal fragment containing the interferon beta gene. J Mol Biol. 204(2):221-31.
2. Hess J, et al., 2004. AP-1 subunits: quarrel and harmony among siblings. J Cell Sci. 117(Pt 25):5965-73.
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