AssayGeneralClass: BAO:0010044 targeted transcriptional assay

AssayMaterialUsed: BTO:0000773 lymphoblastoid cell line derived from control individuals or individuals with germline TP53 variants

AssayDescription: Comparative transcriptomic analysis using RNA-Seq to compare EBV cell lines of wild type and pathogenic TP53 in the context of genotoxic stress induced by doxorubicin treatment. p53 RNA levels were evaluated and expressed as a percentage of the mean levels obtained for the three wild-type TP53 individuals.

AdditionalDocument: PMID: 23172776

AssayReadOutDescription: The p53 mRNA levels were expressed as a ratio of the normal values obtained for 3 TP53 wild-type control individuals.

AssayRange: UO:0000187 the p53 RNA levels were evaluated and expressed as a percentage of the mean levels obtained for three wild-type TP53 individuals.

AssayNormalRange: N/A

AssayAbnormalRange: N/A

AssayIndeterminateRange: N/A

AssayNormalControl: wild type TP53

AssayAbnormalControl: LFS patient cells

ValidationControlPathogenic: 8 Individuals with dominant-negative TP53 missense variants, 10 Individuals with null TP53 variants, and 13 Individuals with other TP53 missense variants

ValidationControlBenign: 3 patients with wild type TP53

Replication: experiments were performed in triplicates.

StatisticalAnalysisDescription: Differentially expressed genes between doxorubicin-treated and untreated cells were arbitrarily defined using, as filters, a P<0.01 and fold-change cutoffs >2 or <2, for up and down regulation, respectively. The resultant signal information was analyzed using one-way analysis of variance (ANOVA, P= 0.001), assuming normality but not equal variances with a Benjamani–Hochberg correction for multiple comparisons using three groups: controls, null, and missense mutations.

SignificanceThreshold: P=0.001

Comment: statistical analysis and P value from previous publication.

AssayGeneralClass: BAO:0010044 targeted transcriptional assay

AssayMaterialUsed: BTO:0000773 lymphoblastoid cell line derived from control individuals or individuals with germline TP53 variants

AssayDescription: Comparative transcriptomic analysis using RNA-Seq to compare EBV cell lines of wild type and pathogenic TP53 in the context of genotoxic stress induced by doxorubicin treatment. 10 biomarkers corresponding to p53 targets were measured to determine a functionality score.

AdditionalDocument: PMID: 23172776

AssayReadOutDescription: In the treated condition, the peak height of each of the 10  p53 target genes was measured and divided by the sum of the heights of the three control genes. This value was then divided by the same ratio calculated in the untreated condition. In the assay, the mean of the 10 values defines the p53 functionality score. The final p53 functionality score is the mean of the scores obtained in RT-MLPA and RT-QMPSF assays.

AssayRange: An arbitrary functionality score was calculated from the induction score of the 10 p53 targets.

AssayNormalRange: N/A

AssayAbnormalRange: N/A

AssayIndeterminateRange: N/A

AssayNormalControl: wild type TP53

AssayAbnormalControl: LFS patient cells

ValidationControlPathogenic: 8 Individuals with dominant-negative TP53 missense variants, 10 Individuals with null TP53 variants, and 13 Individuals with other TP53 missense variants

ValidationControlBenign: 3 patients with wild type TP53

Replication: experiments were performed in triplicates.

StatisticalAnalysisDescription: Differentially expressed genes between doxorubicin-treated and untreated cells were arbitrarily defined using, as filters, a P<0.01 and fold-change cutoffs >2 or <2, for up and down regulation, respectively. The resultant signal information was analyzed using one-way analysis of variance (ANOVA, P= 0.001), assuming normality but not equal variances with a Benjamani–Hochberg correction for multiple comparisons using three groups: controls, null, and missense mutations.

SignificanceThreshold: P=0.001

Comment: statistical analysis and P value from previous publication.