RTPCRRTqPCR

RTPCRRTqPCR is a compounding reference to two related nucleic acid amplification techniques used in molecular biology to study RNA transcripts: reverse transcription PCR (RT-PCR) and real-time quantitative PCR (RT-qPCR). RT-PCR converts RNA into complementary DNA (cDNA) using reverse transcriptase, followed by conventional PCR amplification of target sequences. RT-qPCR, also called real-time RT-PCR, integrates reverse transcription with quantitative PCR, enabling monitoring of amplification in real time through fluorescent signals and allowing estimation of initial RNA amounts.

In RT-qPCR, detection methods include DNA-binding dyes such as SYBR Green or sequence-specific probes (e.g., TaqMan). Quantification can be absolute, using standard curves, or relative, using normalization against stable reference genes and the comparative Ct method (2^-ΔΔCt). Essential controls include no-reverse-transcriptase controls to check for genomic DNA contamination and no-template controls to detect external contamination.

Design considerations involve RNA quality, removal of genomic DNA, primer specificity, and PCR efficiency (ideally near 100%). Endpoint RT-PCR typically uses gel-based or capillary methods to assess product presence, while RT-qPCR yields quantitative data.

Applications span gene expression analysis, pathogen detection, viral load monitoring, and validation of RNA-seq results. Limitations include sensitivity to RNA integrity, potential inhibitors, and the need for careful normalization and interpretation of Ct values.

RTPCRRTqPCR reflects the complementary use of RT-PCR for detection and RT-qPCR for quantification, forming a foundational workflow in molecular diagnostics and research.