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Exome sequencing is a targeted approach that provides a cost-effective alternative to whole genome sequencing for the identification of disease-associated variants, as it targets only the protein coding regions of the human genome. These regions harbor the majority of known disease associated variants. CRS4's human whole exome sequencing service is based on the latest Illumina's high-throughput sequencing platforms and provides a high-quality and convenient solution for the study of complex diseases, rare Mendelian disorders, cancer research and population studies.

Sequencing strategy

• Latest Illumina's HiSeq 3000 systems with 150bp paired-end sequencing

Data quality

• > 85% of bases have a phred quality score ≥ Q30

Turnaround time

• Standard turnaround time is 8 weeks after quality evaluation of the samples

Sample requirements

• The minimum amount of DNA without sample amplification is 5μg in a volume of 20μl.
• DNA must be suspended in TRIS or water – PLEASE DO NOT USE BUFFER CONTAINING EDTA

Data analysis

• SNP and INDEL calling
• Advanced analysis including annotations (allele frequencies in reference datasets - ExAC, 1000Genome, ESP6500; predicted pathogenicity; functional annotations at transcript level with multiple databases - UCSC, Ensembl, RefGene, Gencode) and genetic inheritance
• Additional analysis are available: CNV, extended annotations for non-coding regions
• Custom population scale analysis are available
• Remote access to alignments for visual inspection using IGV
• Additionally raw data are available via secure cloud

Whole genome sequencing provides the most complete picture of the overall genetic variability in an individual. It can be applied, using different sequencing depth, in oncology, to the study of rare disease, and in large scale population studies.

Human whole-genomes are sequenced with the latest Illumina's HiSeq 3000 technology , which makes whole-genome sequencing increasingly cost-effective. Our sequencing platform has successfully sequenced thousands of genomes in the context of two studies on the genetics of autoimmune diseases and longevity.

Sequencing strategy

• Latest Illumina's HiSeq 3000 systems with 150bp paired-end sequencing

Data quality

• > 85% of bases have a phred quality score ≥ Q30

Turnaround time

• Standard turnaround time is 8 weeks after quality evaluation of the samples

Sample requirements

• The minimum amount of DNA without sample amplification is 5μg in a volume of 20μl.
• DNA must be suspended in TRIS or water – PLEASE DO NOT USE BUFFER CONTAINING EDTA

Data analysis

• SNP and INDEL calling
• Advanced analysis including annotations (allele frequencies in reference datasets - ExAC, 1000Genome, ESP6500; predicted pathogenicity; functional annotations at transcript level with multiple databases - UCSC, Ensembl, RefGene, Gencode) and genetic inheritance
• Additional analysis are available: CNV, extended annotations for non-coding regions
• Custom population scale analysis are available
• Remote access to alignments for visual inspection using IGV
• Additionally raw data are available via secure cloud

RNA sequencing (RNA-Seq) is the NGS strategy for the study of the transcriptome, with an higher resolution as compared to other methods such as microarray technology. RNA-Seq based on short reads is widely used to measure the amount of RNA for the identification of differentially expressed genes and pathways in different conditions. Having performed hundreds of RNA-Seq experiments, we have experience with standard RNA-Seq protocols as well as with low-input RNA protocols.

Sequencing strategy

• Latest Illumina systems (HiSeq2500 and HiSeq3000) with 50bp single-end for standard RNA quantification, or longer reads and paired-end sequencing for specific applications (e.g. discovery of novel transcripts)

Data quality

• > 85% of bases have a phred quality score ≥ Q30

Turnaround time

• Standard turnaround time is 4-8 weeks after quality evaluation of the samples

Sample requirements

The RNA used in NGS experiments must be extremely clean in order to get high quality results.

• The minimum amount of RNA without sample amplification is 1μg
• For optimal results, RNA should be treated with DNAse
• The RNA must be resuspended in RNase free water
• Nanodrop measurements, microfluidic electrophoresis RIN or an agarose gel picture should also be provided
• In absence of RIN, the quality of the RNA will be checked at our facility upon reception of the samples
• If low quality is suspected (RIN < 7.5), the procedure will be stopped.
• Proper packaging and cooling of samples is a means to help prevent re-sampling. RNA samples should be delivered at our lab in 2-3 days (e.g. using a courier service) in dry ice.

Data analysis

• Normalized transcript expression
• Identification of differentially expressed genes in different conditions
• Advanced analysis including gene set enrichment analysis for the identification of pathway perturbations
• Remote access to alignments for visual inspection using IGV
• Analysis results are delivered through our web-based platform
• Additionally raw data are available via secure cloud

Whether you are interested in sequencing custom gene panels, libraries or flow-cells, please contact us to discuss your requests.

Custom applications include:

Gene panels: sequencing of large number of samples using gene panels commercially available or designed ad-hoc with your set of genes.

User-made libraries: users can prepare libraries themselves and send them to our lab for sequencing

Illumina flow-cells: users can directly send flow-cells ready to be sequenced