Year-end Sale % OFF 
Abstract
Long non-coding RNA HOX transcript antisense intergenic RNA (HOTAIR) is one of the promising biomarkers that has widely been used in determining the stages of many cancers, including ovarian cancer. In cancer diagnostics, the two key analytical challenges for detecting long non-coding RNA biomarkers are i) the low concentration levels (nM to fM range) in which they are found and ii) the analytical method where broad dynamic range is required (four to six orders of magnitude) due to the large variation in expression levels for different HOTAIR RNAs. To meet these challenges, we report on a biosensing platform for the visual (colorimetric) estimation and subsequent electrochemical quantification of ovarian-cancer-specific HOTAIR using a screen-printed gold electrode (SPE-Au). Our assay utilizes a two-step strategy that involves (i) magnetic isolation and purification of target HOTAIR sequences and (ii) subsequent detection of isolated sequences using a sandwich hybridization coupled with horseradish peroxidase (HRP)-catalyzed reaction of 3,3′,5,5′-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide. The assay achieved a detection limit of 1.0 fM HOTAIR in spiked buffer samples with excellent reproducibility (% RSD ≤ 5%, for n = 3). It was successfully applied to detect HOTAIR in cancer cell lines and a panel of plasma samples derived from patients with ovarian cancer. The analytical performance of the method was validated with standard RT-qPCR. We believe that the proof of concept assay reported here may find potential use in routine clinical settings for the screening of cancer-related lncRNAs.
Highlights
Genome-wide cancer mutation analyses have revealed an extensive landscape of functional mutations within the non-coding genome, with great emphasis on the expression of long non-codingRNAs
Captured HOX transcript antisense intergenic RNA (HOTAIR) targets were magnetically purified with several magnetic washing steps and released from capture probes through heating
The intensity of the colored complex is proportional to the amount of captured horseradish peroxidase (HRP) present in the conjugates, which in turn is proportional to the amount of HOTAIR in the RNA sample
Summary
Genome-wide cancer mutation analyses have revealed an extensive landscape of functional mutations within the non-coding genome, with great emphasis on the expression of long non-codingRNAs (lncRNAs). Long ncRNAs consisting of ∼200 nt or longer transcripts represent a subset of regulatory ncRNAs which are involved in the regulation of post-transcriptional gene expression, epigenomic modulation, and chromatin remodelling, and which have increasingly been identified as key regulators of physiology and pathology [1,2]. HOX transcript antisense intergenic RNA (HOTAIR) is one of the few well-documented lncRNA that is localized on chromosome 12 within the homeobox C (HOXC) gene cluster [6]. This is a 2.2kb-long transcript that is co-expressed with the HOXC gene cluster and influences gene expression via modulation of chromatin dynamics and epigenetic modifications [6].
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
