Method of sequencing a full microrna profile from cerebrospinal fluid
AssigneeThe Translational Genomics Research Institute
InventorsKendall Van Keuren-Jensen, Kasandra L. Burgos
Life of this patent
Estimating the term…
Abstract
The present invention provides methods of purifying RNA from a biological sample comprising two separate aqueous extractions of the organic phase obtained by mixing the biological sample with a first solution comprising guanidinium isothiocyanate and beta-mercaptoethanol and a second solution comprising phenol, chloroform, and isoamyl alcohol. Also provided are methods of sequencing RNA purified from a biological sample and methods of diagnosing Alzheimer's disease and Parkinson's disease in a subject by determining whether a plurality of miRNAs has deregulated expression in biological sample from the subject.
Claims27
A method of purifying RNA from a biological sample, the method comprising:
- a) mixing the biological sample with a first solution comprising guanidinium isothiocyanate and beta-mercaptoethanol to generate a first mixture;
- b) combining a second solution comprising phenol, chloroform, and isoamyl alcohol with the first mixture;
- c) centrifuging the first mixture to produce a first aqueous phase, an interface, and an organic phase;
- d) removing and saving the first aqueous phase;
- e) mixing nuclease-free water with the organic phase and the interface to generate a second mixture;
- f) centrifuging the second mixture to produce a second aqueous phase with the interface and the organic phase;
- g) removing and saving the second aqueous phase with the first aqueous phase;
- and
- h) concentrating and purifying RNA from the first and second aqueous phases with ethanol-based column chromatography or ethanol precipitation and solubilization of the RNA.
A method for diagnosing Alzheimer's disease in a subject, the method comprising:
- a) obtaining a biological sample from the subject;
- b) determining the expression level of a plurality of miRNAs in the biological sample;
- and
- c) detecting Alzheimer's disease in the subject if there is a significant deregulation of the expression levels of the plurality of miRNAs in the biological sample compared to control values.
A method for diagnosing Parkinson's disease in a subject, the method comprising:
- a) obtaining a biological sample from the subject;
- b) determining the expression level of a plurality of miRNAs in the biological sample;
- and
- c) detecting Parkinson's disease in the subject if there is a significant deregulation of the expression levels of the plurality of miRNAs in the biological sample compared to control values.
Dependencies are inferred from the claim text.