Jump to ContentJump to Main Navigation
Show Summary Details

Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter / Tate, Jillian R.


IMPACT FACTOR increased in 2015: 3.017
Rank 5 out of 30 in category Medical Laboratory Technology in the 2014 Thomson Reuters Journal Citation Report/Science Edition

SCImago Journal Rank (SJR) 2015: 0.873
Source Normalized Impact per Paper (SNIP) 2015: 0.982
Impact per Publication (IPP) 2015: 2.238

249,00 € / $374.00 / £187.00*

Online
ISSN
1437-4331
See all formats and pricing

 


Select Volume and Issue
Loading journal volume and issue information...

Evaluation of RNA Isolation Methods and Reference Genes for RT-PCR Analyses of Rare Target RNA

Christine Mannhalter / Daniela Koizar / Gerlinde Mitterbauer

Citation Information: Clinical Chemistry and Laboratory Medicine. Volume 38, Issue 2, Pages 171–177, ISSN (Print) 1434-6621, DOI: 10.1515/CCLM.2000.026, June 2005

Publication History

Published Online:
2005-06-01

Abstract

Reverse transcription polymerase chain reaction (RTPCR) analysis is increasingly becoming part of the diagnostic and prognostic evaluation for hematologic and oncologic disorders. Currently, different RNA isolation methods are used in the diagnostic laboratories. No data are available on their suitability for sensitive detection of breakpoint cluster region-abelson (BCR-ABL) gene transcripts. We have extracted RNA from mononuclear cell (MNC) fractions and from lysed blood samples of 4 patients (1 with leukocytosis, 1 with chronic myelogeneous leukemia (CML) under interferon treatment, and 2 CML patients after bone marrow transplantation) with 3 RNA isolation reagents (TRIzol™, RNAzol™, FastTube™ reagent). RNA yield was slightly higher with RNAzol™ than with TRIzol™ as indicated by agarose gel electrophoresis and spectrophotometric measurement at 260 nm. The FastTube™ reagent was unsuitable for RNA isolation from MNC, and was not evaluated for lysed blood. Quantitative competitive RT-PCR amplification of the ABL gene showed comparable results for RNA isolated with RNAzol™ and TRIzol™. In RNA samples extracted from lysed whole blood, the presence of amplifiable RNA/cDNA was confirmed by amplification of 4 selected reference genes (porphobilinogen deaminase (PBGD), ABL, the gene spanning the BCR on chromosome 22 and retinoic acid receptor alpha (RARA)) in a multiplex PCR. High quality, DNA-free RNA was obtained with RNAzol™, and 1 BCR-ABL-positive (specific for translocation t [9; 22]) cell among 2×104 normal cells was successfully detectable by single step RT-PCR. In RNA isolated with TRIzol™, major contaminations with genomic DNA were observed which significantly impaired the interpretation of the results of RT-PCR analysis.

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

[1]
Ping Wang, Fengxiang Jing, Gang Li, Zhenhua Wu, Zule Cheng, Jishen Zhang, Honglian Zhang, Chunping Jia, Qinghui Jin, Hongju Mao, and Jianlong Zhao
Biosensors and Bioelectronics, 2015
[2]
Kayla A. Chase, Cherise Rosen, Leah H. Rubin, Benjamin Feiner, Anjuli S. Bodapati, Hannah Gin, Edward Hu, and Rajiv P. Sharma
Journal of Psychiatric Research, 2015, Volume 65, Page 87
[3]
Kayla A. Chase, Cherise Rosen, Hannah Gin, Olivia Bjorkquist, Benjamin Feiner, Robert Marvin, Sean Conrin, and Rajiv P. Sharma
Psychiatry Research, 2015, Volume 225, Number 1-2, Page 208
[4]
Eva Sauer, Iris Babion, Burkhard Madea, and Cornelius Courts
Forensic Science International: Genetics, 2014, Volume 13, Page 217
[5]
Eva Sauer, Burkhard Madea, and Cornelius Courts
Forensic Science International: Genetics, 2014, Volume 11, Page 174
[6]
J. Auta, R.C. Smith, E. Dong, P. Tueting, H. Sershen, S. Boules, A. Lajtha, J. Davis, and A. Guidotti
Schizophrenia Research, 2013, Volume 150, Number 1, Page 312
[7]
Yuan Kang, Yi Yin, Qiu Yun Zhang, Lai Sheng Li, Li Xuan Zeng, Ji Wen Luo, and Ming Hung Wong
Environmental Toxicology, 2014, Volume 29, Number 3, Page 354
[8]
Sunghyun Kim, Young Keun Kim, Hyejon Lee, Jang-Eun Cho, Hyo Youl Kim, Young Uh, Young Mi Kim, Hyunjung Kim, Sang-Nae Cho, Bo-Young Jeon, and Hyeyoung Lee
Diagnostic Microbiology and Infectious Disease, 2013, Volume 75, Number 1, Page 68
[9]
E. Mamessier, S. Boniface, P. Dupuy, M. Reynaud-Gaubert, D. Vervloet, and A. Magnan
Journal of Immunological Methods, 2003, Volume 280, Number 1-2, Page 37
[10]
Heather S. Walton, Florian M. Gebhardt, David J. Innes, and Peter R. Dodd
Journal of Neuroscience Methods, 2007, Volume 160, Number 2, Page 294
[11]
F.A. Vlems, T.J.M. Ruers, C.J.A. Punt, Th. Wobbes, and G.N.P. van Muijen
European Journal of Surgical Oncology (EJSO), 2003, Volume 29, Number 4, Page 289
[12]
Yuan Kang, Kwai Chung Cheung, and Ming H. Wong
Toxicology Letters, 2010, Volume 199, Number 3, Page 301
[13]
Margit Mitterbauer, Gerlinde Mitterbauer-Hohendanner, Wolfgang R Sperr, Peter Kalhs, Hildegard T Greinix, Christa Fonatsch, Oskar A Haas, Ulrich Jäger, Christine Mannhalter, and Klaus Lechner
Leukemia & Lymphoma, 2004, Volume 45, Number 5, Page 971
[14]
Michael A. Phillips, John C. D’Auria, Katrin Luck, and Jonathan Gershenzon
Plant Molecular Biology Reporter, 2009, Volume 27, Number 3, Page 407
[15]
Erzsébet Szabó, Éva Korpos, Enkhjargal Batmunkh, Gábor Lotz, Ágnes Holczbauer, Ilona Kovalszky, Ferenc Deák, Ibolya Kiss, Zsuzsa Schaff, and András Kiss
Pathology & Oncology Research, 2008, Volume 14, Number 1, Page 15

Comments (0)

Please log in or register to comment.