Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1998 May 1;26(9):2250–2251. doi: 10.1093/nar/26.9.2250

Highly sensitive detection of gene expression of an intronless gene: amplification of mRNA, but not genomic DNA by nucleic acid sequence based amplification (NASBA).

A Heim 1, I M Grumbach 1, S Zeuke 1, B Top 1
PMCID: PMC147515  PMID: 9547289

Abstract

NASBA is an isothermal nucleic acid amplification reaction that amplifies mRNA in a dsDNA background. Although similar to the sensitive reverse transcription/polymerase chain reaction (RT-PCR) in mRNA detection, NASBA is not prone to give false positive results caused by genomic dsDNA. Therefore, NASBA is unique for sensitive detection of transcription of intronless genes, which preclude strategies such as intron spanning primer pairs to control false positive results in RT-PCR. Using NASBA, mRNA of the intronless human interferon-beta gene was demonstrated with a sensitivity of 10 copies, whereas 100 ng genomic DNA gave a negative result.

Full Text

The Full Text of this article is available as a PDF (46.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Compton J. Nucleic acid sequence-based amplification. Nature. 1991 Mar 7;350(6313):91–92. doi: 10.1038/350091a0. [DOI] [PubMed] [Google Scholar]
  2. Fiers W., Remaut E., Devos R., Cheroutre H., Contreras R., Gheysen D., Degrave W., Stanssens P., Tavernier J., Taya Y. The human fibroblast and human immune interferon genes and their expression in homologous and heterologous cells. Philos Trans R Soc Lond B Biol Sci. 1982 Sep 24;299(1094):29–38. doi: 10.1098/rstb.1982.0103. [DOI] [PubMed] [Google Scholar]
  3. Frielle T., Collins S., Daniel K. W., Caron M. G., Lefkowitz R. J., Kobilka B. K. Cloning of the cDNA for the human beta 1-adrenergic receptor. Proc Natl Acad Sci U S A. 1987 Nov;84(22):7920–7924. doi: 10.1073/pnas.84.22.7920. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fukui H. [Structure of the histamine H1 receptor gene and transcriptional up-regulation of the H1 receptor]. Nihon Rinsho. 1996 Feb;54(2):389–395. [PubMed] [Google Scholar]
  5. Furuta H., Guo D. F., Inagami T. Molecular cloning and sequencing of the gene encoding human angiotensin II type 1 receptor. Biochem Biophys Res Commun. 1992 Feb 28;183(1):8–13. doi: 10.1016/0006-291x(92)91600-u. [DOI] [PubMed] [Google Scholar]
  6. Guatelli J. C., Whitfield K. M., Kwoh D. Y., Barringer K. J., Richman D. D., Gingeras T. R. Isothermal, in vitro amplification of nucleic acids by a multienzyme reaction modeled after retroviral replication. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1874–1878. doi: 10.1073/pnas.87.5.1874. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hattori K., Angel P., Le Beau M. M., Karin M. Structure and chromosomal localization of the functional intronless human JUN protooncogene. Proc Natl Acad Sci U S A. 1988 Dec;85(23):9148–9152. doi: 10.1073/pnas.85.23.9148. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kievits T., van Gemen B., van Strijp D., Schukkink R., Dircks M., Adriaanse H., Malek L., Sooknanan R., Lens P. NASBA isothermal enzymatic in vitro nucleic acid amplification optimized for the diagnosis of HIV-1 infection. J Virol Methods. 1991 Dec;35(3):273–286. doi: 10.1016/0166-0934(91)90069-c. [DOI] [PubMed] [Google Scholar]
  9. Kobilka B. K., Frielle T., Dohlman H. G., Bolanowski M. A., Dixon R. A., Keller P., Caron M. G., Lefkowitz R. J. Delineation of the intronless nature of the genes for the human and hamster beta 2-adrenergic receptor and their putative promoter regions. J Biol Chem. 1987 May 25;262(15):7321–7327. [PubMed] [Google Scholar]
  10. Koike G., Horiuchi M., Yamada T., Szpirer C., Jacob H. J., Dzau V. J. Human type 2 angiotensin II receptor gene: cloned, mapped to the X chromosome, and its mRNA is expressed in the human lung. Biochem Biophys Res Commun. 1994 Sep 30;203(3):1842–1850. doi: 10.1006/bbrc.1994.2402. [DOI] [PubMed] [Google Scholar]
  11. Reis L. F., Ho Lee T., Vilcek J. Tumor necrosis factor acts synergistically with autocrine interferon-beta and increases interferon-beta mRNA levels in human fibroblasts. J Biol Chem. 1989 Oct 5;264(28):16351–16354. [PubMed] [Google Scholar]
  12. van Gemen B., Kievits T., Schukkink R., van Strijp D., Malek L. T., Sooknanan R., Huisman H. G., Lens P. Quantification of HIV-1 RNA in plasma using NASBA during HIV-1 primary infection. J Virol Methods. 1993 Jul;43(2):177–187. doi: 10.1016/0166-0934(93)90075-3. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES