Our high-throughput protocols for Illumina library preparation yield an average success rate of 90%. When possible, we recommend the use of UDIs (unique dual indices) and have a set of 192 for both TruSeq and Nextera applications. For projects requiring higher multiplexing, we can also deploy combinatorial dual indexing for thousands of samples per sequencing run; there is a higher risk of index hopping using combinatorial indexing on patterned flowcells.

DNA sequencing applications

TruSeq (ligation)

• Offering a full coverage of the genome, our TruSeq protocol is optimal when a reference genome is not available. This method can also be used as a PCR-free prep with limited indexing options (up to 12 samples using UDIs per sequencing run, or 96 samples using combinatorial dual indexing).

Nextera (tagmentation) — Skim sequencing

• Skim-sequencing libraries are prepared using a diluted tagmentation enzyme (1/3 concentration). This method is very cost-effective but yields a low coverage. We recommend it for sequencing a very large number of samples for which other genomic resources are available (e.g. haplotype maps). It's not recommended for genomes that do not have a reference genome.

Nextera (tagmentation) — Low volume

• Low-volume libraries are prepared using a smaller reaction volume; the ratio of DNA to tagmentation reagents is maintained. Because the reaction volume is low, you must submit at least 48 samples to have enough final library for sequencing.

Nextera Flex (tagmentation) — Full volume

• Full-volume libraries are prepared using the same method as our lower-throughput Nextera libraries, at lower cost in a plate format.

To be able to process large numbers of samples quickly, we quantify all DNA samples but initially determine size QC for a small subset. If we have concerns about sample quality, we will contact you before proceeding with library preparation. DNA libraries are generated and quantified for each sample, and a custom pool is prepared for sequencing (included in service pricing). Pools for Nextera libraries are also size selected to optimize sequencing output (included). Sequencing is required for all DNA sequencing libraries.

RNA sequencing applications

3’RNAseq

We prepare 3’-tag RNA seq libraries as a cost-effective option for full-transcript RNA-seq. These libraries only capture the 3’ end of mRNAs, with one fragment (read) per transcript.

We highly recommend that you confirm RNA quality prior to submitting samples for 3’RNAseq; determining RNA sample concentrations and quality (integrity) will incur additional charges. Individual 3’RNAseq libraries are generated and quantified for each sample, and a custom pool is prepared for sequencing (included in service pricing). Sequencing is required for all 3’RNAseq libraries.