Transcriptomics, or genome-wide expression profiling, aims to catalogue the complete set of RNA transcripts produced by the genome. Transcriptomics provides new biological insight and can be used to determine the structure of genes, their splicing patterns and other post transcriptional modifications, to detect rare and novel transcripts, and to quantify the changing expression levels of each transcript during development and under different disease conditions.
LC Sciences now provides a one-stop solution (i.e. from sample to data) for transcriptome sequencing using the latest in RNA-Seq technology. Total RNA-Seq is a new method and a powerful tool to identify and quantitatively decode the entire population of RNAs in your sample. Rather than sequencing RNA directly, the RNA-Seq method makes use of the latest high-throughput sequencing technology to sequence cDNA in order to reveal information about your sample’s RNA content.
The Total RNA-Seq (Whole Transcriptome) sequencing services by LC Sciences are comprehensive and provide the most complete picture of RNA content in your samples. We can help you set up and conduct a high-quality, well-controlled Total RNA-Seq experiment based on the latest deep-sequencing technologies.
Transcriptome Profiling Applications
- Detection of mutation events such as: novel splicing events, gene fusions, SNPs, or other specific coding mutations
- Detection of rare, or novel transcripts
- Profiling and comparison of genome-wide expression in different species or otherwise different biological samples
Key Advantages of RNA-Seq
RNA-Seq provides a more comprehensive view of the transcriptome with a single experiment. RNA-Seq enables us to sequence and profile all species of transcripts in your total RNA samples – from mRNA to non-coding RNAs to small RNAs.
RNA-Seq is not necessarily dependent on any prior sequence knowledge. There is no need for design of probes that must be based on prior sequence or secondary structure information. Therefore, transcriptome profiling in any species is possible which makes this method particularly attractive for non-model species. Additionally, RNA-Seq data can be used to build de novo gene models.
RNA-Seq has increased dynamic range and sensitivity. RNA-Seq enables you to achieve “digital” transcript expression analysis meaning that expression level data are based on each individual transcript that is sequenced and counted. By increasing the sequencing depth, a potentially unlimited dynamic range can be reached making RNA-Seq an ideal tool for detection rare transcripts.
RNA-Seq provides information of sequence variation in transcripts. RNA-Seq yields a rich data set including information about post transcriptional mutations and their genomic context. Because RNA-Seq data yields information about how exons are connected, it can reveal sequence variations in the transcripts due to alternative splicing events and provide allele-specific or isoform-specific gene expression information. Additionally, RNA-Seq data is useful for gene mapping functions such as describing the length of UTRs and exon boundaries.
Our Comprehensive Service
LC Sciences provides a transcriptome profiling service using the Illumina high-throughput sequencing technology which enables comprehensive coverage, highly sensitive and specific discovery and profiling of all forms of RNAs in your sample.
This service is comprehensive – from sample to data, providing advanced technology and years of experience in efficient discovery and profiling of RNAs.
Our comprehensive profiling service includes:
- Sample QC
- Sample preparation (ribosomal RNA depletion, adapter attachment, reverse transcription)
- Library preparation (single molecule clonal amplification)
- High-throughput sequencing (RNA-Seq method, Illumina Next-Gen sequencing technology)
- Bioinformatics analysis
- Customer data report
Data Analysis / Results
Bioinformatics package includes:
- Illumina base-calling and analysis
- Raw sequencing data – 100 bp paired-end reads – either FASTA-A or FASTA-Q files with base-calling quality scores
- LC Sciences quality filtered data reduced to mappable reads, a list of unique sequences and their copy numbers
- De novo assembly (if applicable)
- Alignment of RNA-Seq reads to customer specified reference genome
- Identification and construction of splice-junctions
- Report of known transcripts with annotation and abundance
- Report of novel transcript with abundance
- Identification of alternate splicing and report of isoform abundance
- Test for differential expression at gene level and transcript level. (biological replicates required – see Technical Note – Biological Replicate Considerations – Differential Expression Analysis)
- GO and KEGG annotation and enrichment analysis (optional)
- Non-coding RNA information – map to genome, alignment, limited annotation and expression info
- Customer data report – includes a summary of methods and all statistical analysis