ABSTRACT
Tuberculosis (TB) remains a major worldwide health problem and has caused millions of deaths in the past few years. Current diagnostic methods, such as sputum smear microscopy and sputum culture, are time-consuming and cannot prevent the rapid spreading of TB during the diagnostic period. In this connection, detecting biomarkers specific to TB at molecular level in plasma of patients will provide a rapid means for diagnosis. In this study, we first evaluated the differential expression of the long non-coding RNAs (lncRNAs) in the plasma from patients with TB (TB positive), community acquired pneumonia (CAP) and healthy individuals (CG) using lncRNA microarray scanning. It was found that there were 2116 specific lncRNAs differentially expressed in the TB positive samples (1102 up-regulated and 1014 down-regulated), which accounted for 6.96% of total lncRNAs. Twelve differentially expressed lncRNAs discovered in microarray were subsequently validated by using real-time quantitative PCR (RT-qPCR). Two lncRNAs (ENST00000354432 and ENST00000427151) were further validated with more Tuberculosis samples. These results suggested the expression level of lncRNAs and the two validated lncRNAs in plasma could be the potential molecular biomarkers for the rapid diagnosis of Tuberculosis.
Subject(s)
Gene Expression Profiling/methods , Mycobacterium tuberculosis/pathogenicity , Oligonucleotide Array Sequence Analysis , RNA, Long Noncoding/genetics , Tuberculosis, Pulmonary/diagnosis , Case-Control Studies , Diagnosis, Differential , Genetic Markers , Host-Pathogen Interactions , Humans , Mycobacterium tuberculosis/isolation & purification , Predictive Value of Tests , RNA, Long Noncoding/blood , Real-Time Polymerase Chain Reaction , Reproducibility of Results , Sputum/microbiology , Transcriptome , Tuberculosis, Pulmonary/blood , Tuberculosis, Pulmonary/microbiology , WorkflowABSTRACT
The aim of this study was to build a gene chip system with surface plasmon resonance (SPR) technique, for which Gamma-peptide nucleic acid (Gamma-PNA) functioned as a probe, in order to improve sensitivity and its specificity. With the use of self-assembled monolayer (SAM) technology, surface chemistry of two-dimensional structure was used. Gamma-PNA was designed according to the bioinformatics, and was plated on the SPR chip modified by SAM. Subsequently, relevant parameters of the experiment were ensured and optimized. The results showed that the performances of Gamma-PNA probe was little affected by the ion concentration of buffer, and it had a strong light signal in a stable state. As the ion concentration was 0, there were still good hybrid reactions; pH value had less influence upon Gamma-PNA probe, and acid environment of buffer could be better. Gamma-PNA probe combined with sensor technologies achieved made the probe with dispensable labels and real-time detection. It also improved the efficiency of the hybridization and the stability, providing the foundation for clinical application.
