Optimized Expression Profiling for Challenging FFPE Samples
FFPE Expression Profiling
Improve the quality of FFPE expression profiling studies with QuantSeq FFPE 3’ mRNA-Seq gene expression kits. The cost-saving 3’ sequencing approach enables mRNA-focused readout and exceptional scalability to hundreds and thousands of samples.
QuantSeq FFPE generates Illumina-compatible UMI-tagged libraries from total RNA in <4.5 hours without the need for any pre-processing steps and presents the ideal choice for cost-efficient gene expression profiling studies of FFPE samples.
Performance
QuantSeq FFPE 3’ mRNA-Seq is a fast and cost-efficient gene expression profiling kit specifically designed and validated on FFPE samples from various tissues and organisms.
Excellent Performance Across a Wide Range of Input RNA Amounts
The majority of reads generated by QuantSeq FFPE library preparation from 5 ng – 200 ng mouse liver FFPE RNA present uniquely mapping reads (Fig. 1A). As a results, QuantSeq FFPE provides excellent gene detection across a wide range of FFPE input RNA amounts (Fig. 1B).
Figure 1 | Basic Mapping Stats (A) and Gene Detection (B) for QuantSeq FFPE libraries generated from 200 ng, 50 ng, and 5 ng mouse liver FFPE RNA (DV200 = 46.3 %) at 5 M reads per sample.
Robust Performance for Tumor and Matched Normal FFPE Tissues
QuantSeq FFPE generates a high proportion of uniquely mapping reads (up to 79 %) when tested on human kidney tumor and matched normal FFPE tissues (Fig. 2) providing high-quality data for gene expression analysis.
Figure 2 | Basic Mapping Stats for QuantSeq FFPE libraries generated from 20 ng matched human kidney tumor FFPE RNA (DV200 = 45 %) and normal kidney FFPE RNA (DV200 = 50 %).
High Reproducibility for Replicate Samples from Tumor and Normal FFPE Tissues
Gene counts generated with QuantSeq FFPE are highly reproducible between replicates (Fig. 3) indicating robustness and consistency even for variable sample types such as FFPE.
Figure 3 | Gene count correlation analysis for QuantSeq FFPE generated from two replicates using 20 ng matched human kidney tumor FFPE RNA (DV200 = 45 %) and normal kidney FFPE RNA (DV200 = 50 %).
QuantSeq FFPE is Ideal for Differential Gene Expression Analysis from FFPE Samples
QuantSeq FFPE generates one read per transcript from the 3’ end of polyadenylated RNAs using total FFPE RNA as input. QuantSeq FFPE is therefore ideal for expression profiling using straightforward read counting as demonstrated by differential gene expression analysis for human kidney tumor vs. matched normal FFPE RNA (Fig. 4).
Figure 4 | Volcano plot of differentially expressed genes between human kidney tumor and normal FFPE tissue at a padj threshold of <0.01. 20 ng matched human kidney tumor or normal FFPE RNA were inserted into QuantSeq FFPE library preps and sequenced at 5 M reads/sample. Differential expression analysis was performed using DESeq2.
Workflow
QuantSeq FFPE is a simple, all-in-one library prep kit that generates RNA-Seq libraries within 4.5 hours from FFPE and degraded samples with only 1.5 hours hands-on time. The kit uses total RNA as input, hence no prior poly(A) enrichment or rRNA depletion is needed. QuantSeq FFPE is capturing the 3’ end of polyadenylated mRNA and is optimized for low-quality FFPE samples with low input amounts.
FAQ
Frequently Asked Questions
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Ordering Information
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