Identification of MicroRNAs (miRNAs) in Fecal Samples as Biomarkers for Gastroenterological Cancers

A method of detecting a disease in a subject suspected of having the disease comprising the steps of:

• obtaining a stool sample from the subject;
• mixing the stool sample with a solution of a salt, RNase free water or both to form a suspension;
• centrifuging the suspension;
• separating a supernatant from the centrifuged suspension;
• measuring total a RNA/microRNA (miRNA) concentration in the supernatant using a spectrophotometer;
• transcribing cDNA from the total RNA using one or more specific miRNA primers;
• amplifying the transcribed cDNA using a polymerase chain reaction (PCR) assay to obtain one or more amplified miRNAs;
• and
• comparing the levels of the one or more miRNAs from the stool samples of the subject suspected of having the disease with that of one or more healthy subjects, wherein a change in the level of expression of the one or more miRNAs indicates the presence of the disease.

A method of isolating and amplifying one or more microRNAs (miRNAs) from a stool sample comprising the steps of:

• mixing the stool sample with a sodium chloride solution, RNase free water or both to form a suspension, wherein the stool sample to sodium chloride solution ratio is 1:10;
• centrifuging the suspension;
• separating a supernatant from the centrifuged suspension;
• measuring a total RNA/miRNA concentration in the supernatant using a spectrophotometer;
• transcribing a cDNA from the total RNA using one or more specific miRNA primers;
• and
• amplifying the transcribed cDNA using a polymerase chain reaction assay to obtain the one or more amplified miRNAs from the stool sample.

A method of detecting a colorectal or a gastroenterological cancer in one or more subjects comprising the steps of:

• obtaining a stool sample from the one or more subjects suspected of having the colorectal or the gastroenterological cancer;
• mixing the stool sample with a sodium chloride solution, RNase free water or both to form a suspension, wherein a ratio of the stool sample to sodium chloride solution is 1:10;
• centrifuging the suspension;
• separating a supernatant from the centrifuged suspension;
• measuring a total RNA/microRNA (miRNA) concentration in the supernatant using a spectrophotometer;
• transcribing a cDNA from the total RNA using one or more specific miRNA primers;
• amplifying the transcribed cDNA using a polymerase chain reaction assay to obtain one or more amplified miRNAs;
• and
• comparing the levels of the one or more miRNAs from the stool samples of the one or more subjects suspected of having the colorectal or gastroenterological cancer with that of one or more healthy subjects, wherein an elevated level of the one or more miRNAs indicates the presence of the colorectal or gastroenterological cancer.

A method of processing a biological sample for amplification of one more target MicroRNAs (miRNAs) in the sample without a prior extraction of the one or more miRNAs comprising the steps of:

• mixing the biological sample with a salt solution, RNase free water, or both to form a suspension;
• centrifuging the suspension;
• separating a supernatant from the centrifuged suspension;
• and
• storing the supernatant at −80° C. or processing the supernatant immediately to amplify the one or more miRNAs by a polymerase chain reaction.