De Novo RNA Seq:
Identify the genes present in your transcriptome. De novo sequencing of transcriptomes: What is possible? De novo transcriptome sequencing analyses expressed genes present at a specific time and with a specific physiological background. With the average transcript length of 1,500-2,000 bp, several reads have to be generated per transcript, which are later assembled to reconstruct the full length transcript. Sequencing our random primed cDNA library on Illumina NextSeq 500 or HiSeq 2500 provides you with tens of millions of 75 bp, 100 bp or 150 bp reads. Usually the output of one channel is way too high for one transcriptome, meaning that several libraries can be analyzed in one channel. To your convenience, Eurofins has built up a proprietary software pipeline for de novo assembly of short Illumina reads to cost-efficiently reconstruct the transcripts.
- Detect rare or previously unknown transcripts
- Strand specific protocol (upon request)
- Cost-efficient de novo transcriptome data
Reference based RNA Seq:
Analyze gene expression responses and rely on our long-term experience: mRNA-Seq for gene expression analysis and detection of splicing events. Gain one of the most comprehensive views of all expressed transcripts within your sample with our RNA-Seq services. Since the entire transcriptome is covered with short Illumina reads, RNA-Seq is the preferred method to analyze novel transcripts, novel isoforms, alternative splice sites, rare transcripts or SNPs.
- Analyze gene expression, gene expression abundance, alternative splice sites and SNPs.
- Strand-specific protocol (upon request)
- Obtain comprehensive view of the entire transcriptome
- Analyze prokaryotic transcripts by rRNA depletion
We accept isolated RNA, microbial cultures (non‐pathogenic), animal/plant tissues, etc
- RNA: Submit DNase treated RNA of approximately 8-10µg of high quality with Nanodrop A260/280 ratio >1.9; A260/230=2.0-2.2; and at a concentration of ~100 ng/µl (dissolve in nuclease free water). Supplied RNA should be shipped in dry ice to the Eurofins facility.
- Plant/Human/Animal Tissue: Tissues of 2-4 gm should be provided in RNA later or snap frozen tissue without any storage solution shipped in dry ice.
- Microbial non-pathogenic culture: Snap frozen pure isolated culture (Bacteria / Yeast / Fungi) should be shipped in dry ice.
Sample Quality Control:
- NGS grade High quality RNA will be isolated from the supplied sample using commercially available kits.
- The qualification and quantification of RNA will be carried out by denatured agarose gel electrophoresis and NanoDrop/Qubit Fluorometer analysis.
- RIN value estimation will be carried out using Agilent 4200 Tape Station.
- RNA Seq library will be prepared by using TruSeq Stranded mRNA Sample Prep Kit or TruSeq Stranded RNA Library Prep Kit (depending on the requirement).
- Library validation will be carried out using High Sensitivity D1000 Screen tape using Agilent Tape Station.
- Sequencing will be carried out on Illumina NextSeq 500 platforms with 2 x 75 or 2 x 150 bp v2 chemistry or HiSeq 2500 with 2 x125bp v4 chemistry.
- Generation of required amount of data as per the requirement.
- Sequence data (base call files or bcl files) generated from the sequencer are de-multiplexed if required and converted to FASTQ files.
- Raw data will be available for download as a compressed archive of FASTQ files for each sample.
- Raw data can be post processed upon request.
- Comprehensive compiled report and data set in Pen Drive or Hard Disk.
- 5-7 weeks after arrival of your samples and all necessary information. It depends upon data size, scope, technology selected, number of samples and complexity of the project. The TAT is offered expecting no biological or technical difficulties for processing of all project samples.