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Detection of Minimal Residual Disease in Leukaemia by RT-PCR

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PCR Mutation Detection Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 688))

Abstract

Reverse transcription-PCR (RT-PCR) describes a technique whereby RNA is first reverse transcribed into complementary DNA (cDNA) using the enzyme reverse transcriptase, and the resulting cDNA amplified in either a single-step or a two-step nested PCR reaction. It is particularly useful for detecting molecular markers associated with leukaemia, as it enables a single assay to be used for many patients, each with unique genomic translocation breakpoints, but who have common fusion points in mRNA. This molecular method can not only detect aberrations that are cytogenetically cryptic, such as t(12;21)(p13;q22) in paediatric acute lymphoblastic leukaemia (ALL), but is also very sensitive, and as such can be used to monitor minimal residual disease (MRD). Detecting and measuring MRD is important because it can be a guide to determining prognosis and relapse risk, predict recurrence of leukaemia, and enable individualization of treatment.

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References

  1. Mullis, KB and Faloona, FA (1987) Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Methods Enzymol. 155, 335–350.

    Article  PubMed  CAS  Google Scholar 

  2. Cross NCP, Lin F, Chase A, Bungey J, Hughes TP, Goldman JM. (1993) Competitive PCR to estimate the number of BCR–ABL transcripts in chronic myeloid leukemia patients after bone marrow transplantation. Blood 82, 1929–1936.

    PubMed  CAS  Google Scholar 

  3. Hochhaus A, Lin F, Reiter A, Skladny H, Mason PJ, van Rhee F, Shepherd PC, Allan NC, Hehlmann R, Goldman JM, Cross NC (1996) Quantification of residual disease in chronic myelogenous leukemia patients on interferon-alpha therapy by competitive polymerase chain reaction. Blood 87, 1549–1555.

    PubMed  CAS  Google Scholar 

  4. Gabert J, Beillard E, van der Velden VH, Bi W, Grimwade D, Pallisgaard N, Barbany G, Cazzaniga G, Cayuela JM, Cavé H, Pane F, Aerts JL, De Micheli D, Thirion X,Pradel V, González M, Viehmann S, Malec M, Saglio G, van Dongen JJ. (2003) Standardization and quality control studies of ‘real-time’ quantitative reverse transcriptase polymerase chain reaction of fusion gene transcripts for residual disease detection in leukemia – a Europe Against Cancer program. Leukemia 17, 2318–2357.

    Article  PubMed  CAS  Google Scholar 

  5. van Dongen JJ, Macintyre EA, Gabert JA, Delabesse E, Rossi V, Saglio G, Gottardi E, Rambaldi A, Dotti G, Griesinger F, Parreira A, Gameiro P, Diáz MG, Malec M,Langerak AW, San Miguel JF, Biondi A. (1999) Standardized RT-PCR analysis of fusion gene transcripts from chromosome aberrations in acute leukemia for detection of minimal residual disease. Report of the BIOMED-1 Concerted Action: investigation of minimal residual disease in acute leukemia. Leukemia 13, 1901–1928.

    Article  PubMed  Google Scholar 

  6. Hughes T, Deininger M, Hochhaus A, Branford S, Radich J, Kaeda J, Baccarani M, Cortes J, Cross NC, Druker BJ, Gabert J, Grimwade D, Hehlmann R, Kamel-Reid S, Lipton JH, Longtine J, Martinelli G, Saglio G, Soverini S, Stock W, Goldman JM. (2006) Monitoring CML patients responding to treatment with tyrosine kinase inhibitors: review and recommendations for harmonizing current methodology for detecting BCR–ABL transcripts and kinase domain mutations and for expressing results. Blood 108, 28–37.

    Article  PubMed  CAS  Google Scholar 

  7. Thörn I, Olsson-Strömberg U, Ohlsen C, Jonsson AM, Klangby U, Simonsson B, Barbany G. (2005) The impact of RNA stabilization on minimal residual disease assessment in chronic myeloid leukemia. Haematologica 90, 1471–1476.

    PubMed  Google Scholar 

  8. Kwok, S and Higuchi, R (1989) Avoiding false positives with PCR. Nature 339, 237–238.

    Article  PubMed  CAS  Google Scholar 

  9. O’Connell, J (2002) The Basics of RT-PCR, in RT-PCR protocols, O’Connell, J (ed) Humana, Totowa, NJ, pp. 19–25

    Chapter  Google Scholar 

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Mason, J., Griffiths, M. (2011). Detection of Minimal Residual Disease in Leukaemia by RT-PCR. In: Theophilus, B., Rapley, R. (eds) PCR Mutation Detection Protocols. Methods in Molecular Biology, vol 688. Humana Press. https://doi.org/10.1007/978-1-60761-947-5_18

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  • DOI: https://doi.org/10.1007/978-1-60761-947-5_18

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60761-946-8

  • Online ISBN: 978-1-60761-947-5

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