Prelude™ Direct Lysis Module
The Prelude Direct Lysis Module provides a fast and easy method for whole transcriptome analysis using direct cell lysates without RNA isolation. Cell washing and lysis take less than five minutes, and the resulting lysates can be added directly to the first strand synthesis reaction of the Ovation® family of RNA amplification products for microarrays, qPCR and RNA-Seq.
The Prelude Direct Lysis Module provides optimized reagent mixes and a protocol to process 24 samples (Part No. 1400-24).
Fold-Change Correlation of Cell Lysate Versus Purified RNA Samples. The figure shows the concordance of fold-change values for 3,040 genes detected as differentially expressed (p-value <0.001) between lysed HeLa and U937 cells and their purified RNA equivalents. Cell lysates were prepared using the Prelude Direct Lysis Module, and total RNA was isolated from each cell line using the QIAGEN® RNeasy® Mini Kit. Amplification was conducted using the Ovation Pico WTA System. The cDNA product was fragmented and labeled using the Encore™ Biotin Module and hybridized to Affymetrix GeneChip® Human Genome U133A 2.0 Arrays. Differentially expressed genes >2-fold absolute change are shown.
Highly Concordant Results by RNA-Seq. HeLa cells were processed using the Prelude Direct Lysis Module, and total RNA was isolated from HeLa cells using the QIAGEN RNeasy Mini Kit. Cell lysates or isolated total RNA samples were amplified using the Ovation RNA-Seq System, and the resulting double-stranded cDNA used to construct libraries for sequencing on the Illumina® Genome Analyzer IIx platform. Single-read sequencing was done with 40 base-pair reads. Fastq reads were processed with Tophat v1.0.11 and Bowtie 0.12.3 to generate RPKM values (Reads Per Kilobase per Million mapped reads) for 23,581 transcripts. RPKM values above 0.01 were log2 transformed and are plotted as shown.