Sentences with phrase «luciferase reporter»

After 24 hours, luciferase activities were measured by using dual - luciferase reporter assay system (Promega).

Once they'd found suitable pairs, they constructed three components: GBPa - VP16 (activation domain); GBPb - GAL4 (DNA - binding domain); and a UAS - driven luciferase reporter construct.

C and D, luciferase reporter gene assay on the transcription factors for RIG - G gene expression.

The pGL3 luciferase reporter plasmids with the wild - type or mutated 3 ′ UTR of APC were transiently transfected into HEK293T cells along with 25 nM miR - 142 precursor or negative control precursor.

A, schematic representation of serial p21waf1 / cip1 promoter — driven luciferase reporters used in transactivation assay: p21 - Luc contains the 2.3 - kb full - length of p21waf1 / cip1 promoter (from − 2,326 to +10 nucleotide related transcription start site); its derivatives (− 159 / +8) p21 - Luc, (− 123 / +8) p21 - Luc, (− 84 / +8) p21 - Luc, (− 76 / +8) p21 - Luc, (− 63 / +8) p21 - Luc, and (− 56 / +8) p21 - Luc contain a series of deleted promoters of p21waf1 / cip1.

Other p21waf1 / cip1 promoter — driven luciferase reporters containing multiple point mutations of the Sp1 sites or point mutation of the AP - 2 and E2F responsive elements (Mut1 / 2) p21 - Luc, (Mut3 / 4) p21 - Luc, (Mut1 / 2/3) p21 - Luc, (Mut2 / 3/4) p21 - Luc, (Mut1 / 2/3 / 4) p21 - Luc, (mAP -2-1) p21 - Luc, (mAP2 - 2) p21 - Luc, (mE2F (b)-1) p21 - Luc, and (mE2F (b)-2) p21 - Luc were generated using the QuickChange site - directed mutagenesis system (Stratagene, La Jolla, CA).

For DNA transfection, the HepG2, 293, and 293T cells were seeded respectively at 2 × 105, 2 × 105, and 1 × 105 cells / well in 24 - well plates overnight, and then the p53 - TA - Luc luciferase reporter plasmid and RSV - lacZ plasmids were transiently transfected into the cells using NTRII for 6 h, followed by treatment with the test compounds.

Several luciferase reporters with wt or mutant RIG - G promoters were constructed (Fig. 5B and C): pXP2 (− 87) was totally in lack of two ISRE elements; pXP2 - mut - 1 and pXP2 - mut - 2 contained mutations in the site of ISRE I and ISRE II, respectively; and pXP2 - mut - 3 possessed double mutations.

The minor allele of rs25532 significantly decreased luciferase reporter gene expression levels by 15 — 80 %, depending on 5 - HTTLPR allele background and cell type.

The Dual - Luciferase Reporter Assay System (E1910; Promega, Madison, WI) was used to perform dual - reporter assays on NFκB luciferase and Renilla luciferase (internal control).

C, schematic representation of the wt or mutant RIG - G promoter - luciferase reporter gene constructs.

E, constructs carrying KLF4 3 ′ UTR luciferase reporter or deletion mutant of miR7 binding sites (site 1; MT1, site 2; MT2) were transfected in 293TN cells with indicated miRNAs.

Once Chaput's group had acquired a library of 250 distinct translation enhancing elements through selection using mRNA display, the sequences were screened for translation enhancing activity, which was quantified using a light based assay employing a luciferase reporter molecule.

For the luciferase reporter assay, correct amounts of cells (HuH - 7, HeLa, NIH 3T3, 293T, HCT116, and HepG2 cells) were transfected with an appropriate amount of reporter plasmid, p21 - Luc, or its derivatives and either the empty parental vector or the DDX3 expression construct.

The p21waf1 / cip1 promoter — driven luciferase reporters (pGL3 derivatives) containing point mutations in six Sp1 elements, (Mut1) p21 - Luc, (Mut2) p21 - Luc, (Mut3) p21 - Luc, (Mut4) p21 - Luc, and (Mut5 / 6) p21 - Luc and the parental wild - type reporter, (Wt) p21 - Luc, were kindly provided by Dr. D. Kardassis (Department of Basic Sciences, University of Crete Medical School, Heraklion, Crete, Greece; ref.

Next, to examine the underlying mechanism by which DDX3 activates the p21waf1 / cip1 promoter, we mapped the DDX3 responsive region within the p21waf1 / cip1 promoter by serial 5 ′ - deleted promoter - driven luciferase reporters (Fig. 3A and Supplementary Fig.

To examine whether DDX3 transactivates the p21waf1 / cip1 promoter activity through Sp1 site - interacting proteins, the Sp1 or Sp3 expression plasmid together with the DDX3 expression construct and the p21waf1 / cip1 promoter — driven luciferase reporter were introduced into HuH - 7 cells.

p21waf1 / cip1 promoter — driven luciferase reporter (p21 - Luc; 0.05 - 0.5 μg) and increasing amount (0.1 - 2 μg) of FLAG - DDX3 expression construct were cotransfected into various cell lines as indicated.

To this end, increasing amounts of expression constructs of DDX3 / DQAD, DDX3 / AAA and wild - type DDX3 were cotransfected with p21waf1 / cip1 promoter — driven luciferase reporter.

To construct the serial 5 ′ - deleted p21waf1 / cip1 promoter — driven luciferase reporters (− 1,002 / +8) p21 - Luc, (− 463 / +8) p21 - Luc, (− 223 / +8) p21 - Luc, (− 210 / +8) p21 - Luc, (− 159 / +8) p21 - Luc, (− 106 / +8) p21 - Luc, (− 84 / +8) p21 - Luc, (− 76 / +8) p21 - Luc, (− 63 / +8) p21 - Luc, and (− 56 / +8) p21 - Luc, HindIII / BglII — treated, PCR - amplified p21waf1 / cip1 promoter fragments were ligated with HindIII / BglII — treated pGL2 vector.

To more fully explore the effects of compound on Bach1 activity, a luciferase reporter assay using the HMOX1 E2 ARE as a target was developed.

Neural stem cells transfected with a luciferase reporter gene construction were treated with 12 000 compounds.

The clone, pGL - MARE - Luc, was confirmed via DNA sequencing before being used in the luciferase reporter assay.

(a) Luciferase reporter assay of COX - 2 promoter.

Transfection and luciferase reporter assay.

Several IFN signaling pathway — related key regulators, including STAT1, STAT2, IRF - 1, and IRF - 9, were transfected individually or in combination into STAT1 - deficient U3A cells, together with a luciferase reporter gene containing RIG - G promoter.

In agreement with this, both wt STAT2 and mutant STAT2 - Y690F displayed a similar effect on the luciferase reporter gene expression in U3A cells without IFNα.

Luciferase activities were measured using the Dual - Luciferase Reporter Assay System (Promega) and normalized to Renilla luciferase activity.