Background: Next Generation Sequencing technologies have facilitated differential gene expression analysis through RNA-seq and Tag-seq methods. RNA-seq has biases associated with transcript lengths, lacks uniform coverage of regions in mRNA and requires 10-20 times more reads than a typical Tag-seq. Most existing Tag-seq methods either have biases or not high throughput due to use of restriction enzymes or enzymatic manipulation of 5' ends of mRNA or use of RNA ligations.Results: We have developed EXpression Profiling through Randomly Sheared cDNA tag Sequencing (EXPRSS) that employs acoustic waves to randomly shear cDNA and generate sequence tags at a relatively defined position (~150-200 bp) from the 3' end of each mRNA. Implementation of the method was verified through comparative analysis of expression data generated from EXPRSS, NlaIII-DGE and Affymetrix microarray and through qPCR quantification of selected genes. EXPRSS is a strand specific and restriction enzyme independent tag sequencing method that does not require cDNA length-based data transformations. EXPRSS is highly reproducible, is high-throughput and it also reveals alternative polyadenylation and polyadenylated antisense transcripts. It is cost-effective using barcoded multiplexing, avoids the biases of existing SAGE and derivative methods and can reveal polyadenylation position from paired-end sequencing.Conclusions: EXPRSS Tag-seq provides sensitive and reliable gene expression data and enables high-throughput expression profiling with relatively simple downstream analysis. © 2014 Rallapalli et al.; licensee BioMed Central Ltd.
CITATION STYLE
Rallapalli, G., Kemen, E. M., Robert-Seilaniantz, A., Segonzac, C., Etherington, G. J., Sohn, K. H., … Jones, J. D. G. (2014). EXPRSS: An Illumina based high-throughput expression-profiling method to reveal transcriptional dynamics. BMC Genomics, 15(1). https://doi.org/10.1186/1471-2164-15-341
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