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Single Monochrome Real-Time RT-PCR Assay for Identification, Quantification, and Breakpoint Cluster Region Determination of t(9;22) Transcripts  Marina.

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Presentation on theme: "Single Monochrome Real-Time RT-PCR Assay for Identification, Quantification, and Breakpoint Cluster Region Determination of t(9;22) Transcripts  Marina."— Presentation transcript:

1 Single Monochrome Real-Time RT-PCR Assay for Identification, Quantification, and Breakpoint Cluster Region Determination of t(9;22) Transcripts  Marina I. Gutiérrez, Georgina Timson, Abdul K. Siraj, Rong Bu, Shakuntala Barbhaya, Sripad Banavali, Kishor Bhatia  The Journal of Molecular Diagnostics  Volume 7, Issue 1, Pages (February 2005) DOI: /S (10) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

2 Figure 1 The top graph represents the involved area of the BCR and ABL genes with the indicated location of primers in e1, b2 and a2. Exons are numbered. A: Melting curve analysis for identification of various BCR-ABL transcripts. Discrete melting peak temperatures are obtained for each fusion as indicated. Dotted lines represent amplicons from cDNA from the control cell lines, K562 and SUPB15. The other peaks correspond to patient samples. B: Conventional RT-PCR analysis demonstrates different size bands. Lanes: M is a 100-bp marker and the rest are indicated by the BCR exon (b3, e1, or b2) joint to ABL exon 2 from independent patient samples. The Journal of Molecular Diagnostics 2005 7, 40-47DOI: ( /S (10) ) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

3 Figure 2 Chromatograms obtained by direct sequencing of PCR products, indicating the junction b3-a2 (two samples) and b2-a2 (two samples). Breakpoints (bkpt) are indicated. The Journal of Molecular Diagnostics 2005 7, 40-47DOI: ( /S (10) ) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

4 Figure 3 Standard curves for quantification of number of transcripts using serial 10-fold dilutions of a plasmid containing either e1-a2 (left) or b3-a2 (right) junctions. Copy numbers are indicated for the highest and lowest concentrations. The Journal of Molecular Diagnostics 2005 7, 40-47DOI: ( /S (10) ) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

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