Characterization
Description(s):NFkB is a ubiquitous transcription factor that plays a key role in cellular responses to numerous stimuli such as stress, cytokines, ultraviolet irradiation, and bacterial or viral antigens. Incorrect regulation of NFkB has been linked to cancer, inflammatory and autoimmune diseases, and septic shock. When NFkB is activated, it dissociates from its inhibitor IkB and moves from the cytoplasm to the nucleus. Once in the nucleus, it binds to target DNA elements and positively regulates the transcription of genes involved in immune and inflammatory responses, cell growth control, and apoptosis.
Signosis has developed a plate-based array for profiling 20+ NFkB-regulated genes. By using this assay, researchers are able to compare gene expression in up to three samples on a single microtiter plate.
Applicable Grid:
List of Applicable Genes
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | |
| A | BCL2 | IL6 | Myc | BCL2 | IL6 | Myc | BCL2 | IL6 | Myc | BCL2 | IL6 | Myc |
| B | 18S rRNA | IL9 | NOS2 | 18S rRNA | IL9 | NOS2 | 18S rRNA | IL9 | NOS2 | 18S rRNA | IL9 | NOS2 |
| C | CCND1 | GAPDH | p53 | CCND1 | GAPDH | p53 | CCND1 | GAPDH | p53 | CCND1 | GAPDH | p53 |
| D | Cox2 | IFNg | TNFA | Cox2 | IFNg | TNFA | Cox2 | IFNg | TNFA | Cox2 | IFNg | TNFA |
| E | FASL | IRF1 | TNFR | FASL | IRF1 | TNFR | FASL | IRF1 | TNFR | FASL | IRF1 | TNFR |
| F | INFB1 | MDR1(Abcb1b) | VCAM1 | INFB1 | MDR1(Abcb1b) | VCAM1 | INFB1 | MDR1(Abcb1b) | VCAM1 | INFB1 | MDR1(Abcb1b) | VCAM1 |
| G | IL1A | MMP1 | VEGFC | IL1A | MMP1 | VEGFC | IL1A | MMP1 | VEGFC | IL1A | MMP1 | VEGFC |
| H | IL2 | Myb | Blank | IL2 | Myb | Blank | IL2 | Myb | Blank | IL2 | Myb | Blank |
Principle
Signosis’ proprietary cDNA plate array is a plate-based hybridization profiling analysis for monitoring the expression of dozens of genes through reverse transcription of mRNA into cDNA. Like array analyses, total RNA is first reverse transcribed into cDNA in the presence of biotin-dUTP in the assay. Targeted genes are then specifically captured onto individual wells on a plate, instead of membranes, through a pre-coated gene-specific oligonucleotide. The captured cDNAs are further detected with streptavidin-HRP. Luminescence is reported as relative light units (RLUs) on a microplate luminometer. The expression level of genes is directly proportional to the luminescent intensity.
Data
Mouse NFkB-regulated cDNA plate array analysis. NIH3T3 cells were treated with (red bars) or without (light blue) 10ng/ml TNF for 30 min. Cells were collected for profiling 23 genes with the cDNA plate array assay.
