Comparative detection and quantitation of human CDK inhibitor mRNA expression of p15INK4B, p16INK4A, p16beta, p18INK4C, p19INK4D, p21WAF1, p27KIP1 and p57KIP2 by RT-PCR using a polycompetitive internal standard.
For comparative and quantitative analysis of human cyclin-dependent kinase inhibitor gene expression (CKI; p15INK4B, p16INK4A, p16beta, p18INK4C, p19INK4D, p21WAF1, p27KIP1 and p57KIP2) we set up an RT-PCR assay with a construct termed pCKIquant producing polycompetitive RNA as an internal standard. We demonstrated the reproducibility, accuracy and high sensitivity of the assay in the in vitro model of myeloid leukaemic HL-60 cells. We also showed that the pCKIquant CKI assay is an excellent tool for the assessment of CKI mRNA expression in clinical samples, e.g. single cryostat sections of lymphoma biopsies.
Schwaller
J
J
Department for Clinical Research, University of Berne, Switzerland.
Pabst
T
T
Bickel
M
M
Borisch
B
B
Fey
M F
MF
Tobler
A
A
D015152:Blotting, Northern; D002352:Carrier Proteins / Q000032:analysis / Q000378:metabolism; D018797:Cell Cycle Proteins; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis; D050763:Cyclin-Dependent Kinase Inhibitor p18; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D050761:Cyclin-Dependent Kinase Inhibitor p57; D018844:Cyclin-Dependent Kinases / Q000032:analysis*; D016213:Cyclins / Q000032:analysis; D004791:Enzyme Inhibitors / Q000032:analysis*; D018922:HL-60 Cells / Q000378:metabolism; D006801:Humans; D007951:Leukemia, Myeloid / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000032:analysis; D009687:Nuclear Proteins / Q000032:analysis; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012333:RNA, Messenger / Q000378:metabolism*; D012680:Sensitivity and Specificity; D025521:Tumor Suppressor Proteins
C493956:CDKN1A protein, human; C493964:CDKN1C protein, human; C493966:CDKN2B protein, human; C493971:CDKN2C protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D050762:Cyclin-Dependent Kinase Inhibitor p15; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050763:Cyclin-Dependent Kinase Inhibitor p18; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050761:Cyclin-Dependent Kinase Inhibitor p57; D016213:Cyclins; D004791:Enzyme Inhibitors; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases