Description:
p53playsacrucialroleinmaintainingorderedproliferation,growth,anddifferentiationofnormalcells.Whilep53normallyexistsinthecellataverylowlevelandinarelativelyinactiveform,inresponsetocellularstressitaccumulatesinthenucleus,whereitbindstospecificDNAsequences.p53isknowntoinduceorinhibittheexpressionofmorethan100targetgenes.Mutationsofthep53geneisthemostfrequentlyencounteredgeneticeventincancer,therebyinactivatingp53andresultinginthedisruptionoftheexpressionoftargetgenes.Therefore,expressionprofilingofp53targetgeneswillfacilitatethefunctionalstudyofp53.ApplicableGrid:
ListofApplicableGenes
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | |
| A | 14-3-3 | CyclinB | MSH2 | 14-3-3 | CyclinB | MSH2 | 14-3-3 | CyclinB | MSH2 | 14-3-3 | CyclinB | MSH2 |
| B | B-actin | GADD45 | p21 | B-actin | GADD45 | p21 | B-actin | GADD45 | p21 | B-actin | GADD45 | p21 |
| C | Bax | HNF4A | PTEN | Bax | HNF4A | PTEN | Bax | HNF4A | PTEN | Bax | HNF4A | PTEN |
| D | BCL-2 | IGF-2 | RB1 | BCL-2 | IGF-2 | RB1 | BCL-2 | IGF-2 | RB1 | BCL-2 | IGF-2 | RB1 |
| E | cdc2 | Jun | Stat3 | cdc2 | Jun | Stat3 | cdc2 | Jun | Stat3 | cdc2 | Jun | Stat3 |
| F | cdc25 | MDM2 | TGFbeta | cdc25 | MDM2 | TGFbeta | cdc25 | MDM2 | TGFbeta | cdc25 | MDM2 | TGFbeta |
| G | CDKN2A | MDR1 | TIMP-3 | CDKN2A | MDR1 | TIMP-3 | CDKN2A | MDR1 | TIMP-3 | CDKN2A | MDR1 | TIMP-3 |
| H | c-myc | MMP1 | VEGF | c-myc | MMP1 | VEGF | c-myc | MMP1 | VEGF | c-myc | MMP1 | VEGF |
Principle:
Signosis’proprietaryCDNAplatearrayisaplate-basedhybridizationprofilinganalysisformonitoringtheexpressionofdozensofgenesthroughreversetranscriptionofmRNAintocDNA.Likearrayanalyses,totalRNAisfirstreversetranscribedintocDNAinthepresenceofbiotin-dUTPintheassay.Targetedgenesarethenspecificallycapturedontoindividualwellsonaplate,insteadofmembranes,throughapre-coatedgene-specificoligonucleotide.ThecapturedcDNAsarefurtherdetectedwithstreptavidin-HRP.Luminescenceisreportedasrelativelightunits(RLUs)onamicroplateluminometer.Theexpressionlevelofgenesisdirectlyproportionaltotheluminescentintensity. |
Data:
Humanp53-regulatedcDNAplatearrayanalysis. TotalRNAsfrom293andHelawereprepared.RNAsweresubjectedtop53-regulatedcDNAplateassay,anddetectedwithchemiluminencebyaplatereader
