1. Quantitative analysis of multiple genes' expressions based on a novel competitive RT-PCR assay.
- Author
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Li, Jia, Lin, Li-hui, Wang, Juan, Peng, Xia, Liu, Ya-nan, Xiao, Jun-hua, Zhou, Yu-xun, and Li, Li
- Subjects
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GENE expression , *DNA primers , *POLYMERASE chain reaction , *SEPARATION (Technology) , *INTERLEUKIN-4 , *INTERLEUKIN-13 , *CONTACT dermatitis , *CELLULAR signal transduction , *PATIENTS - Abstract
We established a novel gene expression analysis platform, Multiplex Competitive RT-PCR Using Fluorescent Universal Primers (MCF-PCR), to study multi-gene expression patterns simultaneously. This platform combines fluorescent universal primers, multiplex competitive RT-PCR, and capillary electrophoretic separation, which ensures MCF-PCR a reliable, medium-throughput, cost-effective technology for gene expression profiling. With cloned standard DNAs, the detection limits, precision, and sensitivity of MCF-PCR were evaluated and compared with that of the assay without adding competitive templates and real-time PCR, respectively. The results showed that detection limit was 3.125 × 10 to 3.2 × 10 copies, and 10 % copy differences between two samples can be detected by MCF-PCR. To validate MCF-PCR, we analyzed expression profile of five genes in interleukin (IL)-4/IL-13 pathway in peripheral blood of 20 healthy adults and 20 allergic dermatitis patients; three genes including IL-4, IL-13, and STAT6 were found differentially expressed in the two sample groups, which maybe key players in IL-4/IL-13 immunological signaling pathway and need further function analysis. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]
- Published
- 2013
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