U6 is not a suitable endogenous control for the quantification of circulating microRNAs

Area of Science:

• Biochemistry
• Molecular Biology
• Genomics

Background:

• Circulating microRNAs (miRNAs) are promising biomarkers for various diseases.
• Their stability in blood makes them ideal for diagnostic applications.
• A lack of reliable internal reference genes hinders circulating miRNA research.

Purpose of the Study:

• To evaluate the suitability of U6 as an internal reference gene for circulating miRNAs.
• To compare the stability of U6 with other miRNAs (miR-16, miR-24) in serum samples.

Main Methods:

• Serum miRNA profiling using microarrays in nasopharyngeal carcinoma patients and controls.
• Validation using real-time quantitative Polymerase Chain Reaction (qPCR) in cancer patients and controls.
• Assessment of miRNA stability under repeated freeze-thaw cycles.

Main Results:

• Microarray analysis indicated differential expression of several miRNAs, excluding U6, between cancer patients and controls.
• U6 exhibited significant expression fluctuations across 110 serum samples.
• U6 expression decreased after freeze-thaw cycles, while miR-16 and miR-24 remained stable.

Conclusions:

• U6 is unsuitable as an internal reference gene for circulating miRNA studies.
• miR-16 and miR-24 demonstrate greater stability and are potential candidates for endogenous controls.