Using Multiple Techniques for Gene Expression Analysis

Reverse Transcription Quantitative PCR
(RT-qPCR)
Custom primer pairs target genes of interest, and fluorescence from reporter molecules measured over time indicates gene expression. While RT-qPCR is the most common and cost-effective approach, it is semi-quantitative, and PCR inhibitors can skew results.

Digital PCR (dPCR)
Each PCR reaction is partitioned into thousands of nanoreactions, which allows this method to detect low-abundance targets. dPCR also tolerates common PCR inhibitors and provides absolute quantification of nucleic acids without the need for standard curves.

RNA Sequencing (RNA-seq)
This method sequences all of the RNA transcripts in a sample, even if certain gene sequences are unknown. RNA-seq provides a comprehensive and unbiased view of the transcripts but is costly and requires expertise to analyze.

Step 1: RNA-seq

An RNA-seq approach allows you to characterize many genes at once, and new advancements make sequencing at the tissue or single-cell level possible.

Step 2: RT-qPCR or dPCR

Validate your RNA-seq results with PCR-based approaches, which are faster, easier, and more affordable than RNA-seq.