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In: Biology

Describe the range of techniques that can be used to monitor human gene expression. Compare high-resolution...

Describe the range of techniques that can be used to monitor human gene expression. Compare high-resolution techniques, used to analyze individual genes and high-throughput techniques that analyze all human genes. (MIn 2 and a half pages)

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1) DNA microarray-

  • Microarray- based studies are uncovering broad patterns of genetic activity, providing new understanding of gene functions and transcriptional processe as well as biological mechanisms.
  • Microarray technology is a powerful technique used to compare differences in gene expression between two mRNA samples.
  • Comparing RNA prepared from diseased cells and normal cells can lead to the identification of sets of genes that play key roles in diseases.
  • Genes that are overexpressed or underexpressed in the diseased cells often present excellent targets for therapeutic drugs. The process uses microarray chips, prepared commercially, which comprise numerous wells, each of which contains an isolated gene.
  • mRNA is extracted from the ‘normal’ sample, and a fluorescent labeled cDNA probe is generated, representing all of the genes expressed in the reference sample.
  • A second cDNA probe is generated using a different-colored fluorescent label and mRNA extracted from the ‘affected’ cells.

2) Quantitative real time rt PCR-

  • Two-step quantitative real-time RT-PCR (RT-qPCR), also known as real-time RT-PCR, kinetic RT-PCR, or quantitative fluorescent RT-PCR, has become the method of choice for gene expression.
  • It is a fast and convenient PCR method that combines traditional RT-PCR with the phenomenon of fluorescence resonance energy transfer (FRET) using fluorogenic primers.
  • The detection of changes in fluorescence intensity during the reaction enables the user to follow the PCR reaction in real time.

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