Product Name Catalog # Price   Qty
Promoter-Binding TF Profiling Array II FA-2002 $999
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Characterization

Description:

To characterize transcription factors (TFs) that binds to a specific promoter or that regulate the expression of a specific gene via its upstream promoter, two common approaches are applied. First is to employ gel shift assay with DNA binding sites of TFs that are silico-identified within the promoter. Second is to removal or knockout the binding site(s) of a specific TF in order to measure whether the expression of a promoter-linked reporter is increased or decreased. Often, a series of reporter constructs with the promoter deletions or mutations need to make because many binding sites of one or a few TFs are present within a promoter. Signosis has developed a fast method to facilitate the characterization of promoters through a revised TF activation array. This assay will help to test whether a selected 96 TFs bind to the promoter or not.


Applicable Grid:

List of Applicable TFs:

   1  2  3

 4

 5

 6

 7  8  9  10

 11

 12
A AP1 CDP GATA

NF-1

Pit

Stat3

XBP FOXG1 HoxA-5 NRF2(A)

ProX1

SOX2 
B AP2 CREB GR/PR NFAT PPAR Stat4  AP3  FOXO1  HSF Oct-1 RB SOX9 
C AR E2F-1

HIF

NF-E2 PXR Stat5 AP4  FREAC-2  KLF4 Pax2 RUNX SOX18
D ATF2 EGR HNF4 NFkB SMAD Stat6 COUP-TF  Gfi-1  MyoD Pax3 ROR(RZR) TFE3 
E Brn-3 ER IRF OCT4 Sp1 TCF/LEF ELK Gli-1 MZF Pax8 RXR TFEB
F C\EBP Ets MEF2 p53 SRF TR FOXA1 HEN (NSCL-1) Nkx2-5 PIT1 SF-1 USF-1
G CAR FAST-1 Myb Pax-5 SATB1 YY1 FOXC1 HNF-1 Nkx3-2 PLAG1

SMUC

VDR
H CBF GAS/ISRE Myc-Max Pbx1 Stat1 TFIID FOXD3  HOX4C NRF1 MEF1 Snail  WT1

 


Principle

Promoter-binding TF profiling plate arrays are a competition assay of Signosis’ TF activation plate arrays. In the TF activation plate arrays, if all of 48 or 96 targeted transcription factors exist in the assayed samples, they will form 48 or 96 types of complexes, each TF with its corresponding biotin-labeled oligo (just like the complex in the gel shift assay). After a simple spin separation of the complexes from unbound free biotin-labeled oligos with a membrane-based column, TF-bound oligos eluted from the column and used for plate hybridization in which a complementary DNA of biotin-labeled oligo. The captured oligo is then detected with streptavidin-HRP and a chemiluminescent substrate. If any TF is not present, it will not form a complex, leading to no detection of TF in the following plate assay of biotin-labeled oligo. In promoter-binding TF profiling arrays, PCR fragment containing the promoter of your interest is mixed with a set of 48 or 96 biotin-labeled oligos corresponding to 48 or 96 TFs along with an assayed sample. If unlabeled promoter DNA fragment contains a TF binding sequence, it will compete with the biotin-labeled oligo to bind to the TF in the sample, leading to no or less biotin labeled TF/DNA complex formation and no or lower detection. Through comparison in the presence and absence of the competitor promoter DNA fragment, promoter-bound TFs can be identified.


Data



Promoter-Binding TF Profiling Assay: HeLa cells were treated with or without PMA. PMA was used to acticve TFs including AP1 and NFkB. Nuclear extracts were prepared and incubated with TF binding oligo probe mix: control HeLa cells without PMA treatment with the probe mix (blue); PMA-treated HeLa cells with the probe mix alone (red) and the probe mix plus IL6 promoter DNA fragment (yellow).

Literature

View user manual

Citations

  1. Cross-talk between macrophages and atrial myocytes in atrial fibrillation. Zewei SunDongchen ZhouXudong XieShuai WangZhen WangWenting ZhaoHongfei XuLiangrong Zheng. Basic Research in Cardiology, 2016. DOI: 10.1007/s00395-016-0584-z
  2. Snail-activated long non-coding RNA PCA3 up-regulates PRKD3 expression by miR-1261 sponging, thereby promotes invasion and migration of prostate cancer cells. JH He, BX Li, ZP Han, MX Zou, L Wang, YB Lv. Tumor Biology, 2016 Original Article First Online: 14 October 2016. DOI: 10.1007/s13277-016-5450-y
  3.  Myocyte enhancer factor (MEF)-2 plays essential roles in T-cell transformation associated with HTLV-1 infection by stabilizing complex between Tax and CREB. P Jain, A Lavorgna, M Sehgal, L Gao, R Ginwala.  Retrovirology, 2015Jain et al. Retrovirology(2015) 12:23 DOI 10.1186/s12977-015-0140-1
  4.  Functional and Activity Analysis of Cattle UCP3 Promoter with MRFs-Related Factors. Chen, W., Xu, H., Chen, X., Liu, Z., Zhang, W., & Xia, D. (2016). IJMS International Journal of Molecular Sciences, 17(5), 682. doi:10.3390/ijms17050682
  5.  Up-Regulation of CREG Expression by the Transcription Factor GATA1 Inhibits High Glucose- and High Palmitate-Induced Apoptosis in Human Umbilical Vein Endothelial Cells. Liu, Y., Tian, X., Li, Y., Liu, D., Liu, M., Zhang, X., Han, Y. (2016). PLOS ONE PLoS ONE,11(5). doi:10.1371/journal.pone.0154861
  6. Characterization of the NLRC5 promoter in chicken: SNPs, regulatory elements and CpG islands. XM Guo, XP Liu, GB Chang, L Xu, YL Bi, HZ Wang DOI: 10.1111/age.12450 Animal Genetics, 2016
  7.  C3 exoenzyme impairs cell proliferation and apoptosis by altering the activity of transcription factors
    L von Elsner, S Hagemann, I Just, A Rohrbeck DOI: 10.1007/s00210-016-1270-2 Naunyn-Schmiedeberg's Archives of Pharmacology 2016
  8.  MGMT DNA repair gene promoter/enhancer haplotypes alter transcription factor binding and gene expression
    M Xu, CE Cross, JT Speidel, SZ Abdel-Rahman doi:10.​1007/​s13402-016-0286-4 Cellular Oncology, 2016
  9. Overexpression of Long Non-coding RNA MEG3 Inhibits Proliferation of Hepatocellular Carcinoma Huh7 Cells via Negative Modulation of miRNA-664. Jin-Hua He, Ze-Ping Han, Ji-Ming Liu, Jia-Bin Zhou, Mao-Xian Zou, Yu Bing Lv, Yu-Guang Li, Ming-Rong Cao. Journal of Cellular Biochemistry. 2017 April 4. doi: 10.1002/jcb.26018.