Ratio-Based Method To Identify True Biomarkers by Normalizing Circulating ncRNA Sequencing and Quantitative PCR Data.

Area of Science:

• Biochemistry
• Molecular Biology
• Genomics

Background:

• Circulating noncoding RNAs (ncRNAs) show promise as disease biomarkers.
• Low concentrations in plasma/serum pose normalization challenges for sequencing and RT-qPCR.
• Existing normalization methods using spiked-in or endogenous controls are unreliable.

Purpose of the Study:

• To address the normalization difficulties in plasma/serum ncRNA analysis.
• To develop a robust method for identifying differentially expressed ncRNAs as biomarkers.
• To improve the reliability of ncRNA biomarker discovery for human diseases.

Main Methods:

• Investigated a ratio-based normalization method using pairs of ncRNAs within the same sample.
• Mathematically verified the independence of this method from external controls.
• Compared the robustness of the ratio-based method against existing normalization techniques.

Main Results:

• The ratio-based method is independent of spiked-in and internal controls.
• This approach is more robust than current reference control-based normalization methods.
• The method reliably identifies differentially expressed ncRNAs as potential disease biomarkers.

Conclusions:

• The ratio-based normalization method effectively solves the normalization problem for circulating ncRNA data.
• This technique enables the reliable identification of ncRNA biomarkers for clinical applications.
• The findings facilitate the use of ncRNAs in disease diagnosis and prognosis.