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Quantitative Analysis of Gene Expression Relative to 18S rRNA in Carcinoma Samples Using the LightCycler® Instrument and a SYBR GreenI-based Assay: Determining FAT10 mRNA Levels in Hepatocellular Carcinoma

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2008

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Lukasiak, Sebastian
Breuhahn, Kai
Schiller, Claudia

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MARX, Andreas, ed., Oliver SEITZ, ed.. Molecular beacons : signalling nucleic acid probes, methods, and protocols. Totowa, NJ: Humana Press, 2008, pp. 59-72. Methods in molecular biology. 429. ISBN 978-1-58829-700-6

Zusammenfassung

Due to the fact that mutations and up- or downregulation of genes can lead to the development of cancer, quantitative comparison of relative gene expression in healthy and cancerous tissue can gain valuable insights into tumorigenesis. While the semi-quantitative DNA microarrays are being used to identify differentially expressed genes on a genomic scale, real-time RT-PCR provides a powerful tool for quantitative measurement of gene expression. Presently, it is the most sensitive method available. Here we describe in detail a SYBR GreenI-based assay using the LightCycler® instrument to measure the levels of mRNA for the ubiquitin-like protein FAT10 relative to 18S rRNA in human hepatocellular carcinoma tissue. This method can be easily adapted to any tissue (human or mouse, rat, etc.) and any gene.

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Fachgebiet (DDC)
570 Biowissenschaften, Biologie

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Relative gene expression, quantitative RT-PCR, SYBR GreenI, co-application reverse transcription (Co-RT), normalization to 18S rRNA

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ISO 690LUKASIAK, Sebastian, Kai BREUHAHN, Claudia SCHILLER, Gunter SCHMIDTKE, Marcus GRÖTTRUP, 2008. Quantitative Analysis of Gene Expression Relative to 18S rRNA in Carcinoma Samples Using the LightCycler® Instrument and a SYBR GreenI-based Assay: Determining FAT10 mRNA Levels in Hepatocellular Carcinoma. In: MARX, Andreas, ed., Oliver SEITZ, ed.. Molecular beacons : signalling nucleic acid probes, methods, and protocols. Totowa, NJ: Humana Press, 2008, pp. 59-72. Methods in molecular biology. 429. ISBN 978-1-58829-700-6
BibTex
@incollection{Lukasiak2008Quant-1270,
  year={2008},
  title={Quantitative Analysis of Gene Expression Relative to 18S rRNA in Carcinoma Samples Using the LightCycler® Instrument and a SYBR GreenI-based Assay: Determining FAT10 mRNA Levels in Hepatocellular Carcinoma},
  number={429},
  isbn={978-1-58829-700-6},
  publisher={Humana Press},
  address={Totowa, NJ},
  series={Methods in molecular biology},
  booktitle={Molecular beacons : signalling nucleic acid probes, methods, and protocols},
  pages={59--72},
  editor={Marx, Andreas and Seitz, Oliver},
  author={Lukasiak, Sebastian and Breuhahn, Kai and Schiller, Claudia and Schmidtke, Gunter and Gröttrup, Marcus}
}
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