Design and application of 60mer oligonucleotide microarray in SARS coronavirus detection

Area of Science:

• Molecular Biology
• Virology
• Biotechnology

Background:

• Severe Acute Respiratory Syndrome (SARS) is a serious respiratory illness caused by a specific coronavirus.
• Accurate and rapid diagnostic methods are crucial for controlling infectious disease outbreaks.
• Existing diagnostic techniques may have limitations in sensitivity or throughput.

Purpose of the Study:

• To design and validate a 60-mer oligonucleotide microarray for the detection of SARS coronavirus.
• To assess the microarray's diagnostic capability and potential for improving detection rates.
• To explore its utility in monitoring viral gene expression during different disease stages.

Main Methods:

• Design of 30 specific 60-mer oligonucleotide probes covering the entire SARS-CoV genome (TOR2 strain).
• Synthesis and printing of probes onto a 12x12 microarray.
• RNA extraction from patient samples (throat swabs, gargling fluid), followed by cDNA synthesis.
• Restriction Display (RD) technique for cDNA fragmentation, labeling with Cy5-universal primer, and microarray hybridization.
• Washing, scanning, and analysis of hybridization signals.

Main Results:

• SARS patient samples showed specific hybridization signals with multiple SARS probes on the microarray.
• Negative and blank controls exhibited no signal, confirming assay specificity.
• The microarray demonstrated parallel detection of the SARS coronavirus genome.
• Improved positive diagnostic ratios were observed compared to unspecified conventional methods.

Conclusions:

• The 60-mer oligonucleotide microarray is a viable tool for parallel detection of the SARS coronavirus genome.
• This microarray technology offers potential for enhanced diagnostic sensitivity and efficiency.
• The platform can be utilized for monitoring SARS-CoV gene dynamics throughout disease progression.