High Throughput DNA Sequencing
We have Illumina HiSeq 2500, NextSeq 500, and MiSeq sequencers that can be used for a broad range of applications, including but not limited to:
- Whole genome sequencing
- Exome sequencing
- Targeted sequencing
- RNA sequencing
- Protein-DNA/RNA interactions (ChIP-Seq, CLIP, etc.)
- Targeted sequencing
- Small RNA sequencing
- ATAC sequencing
- Ribosomal profiling
Service and Sample Submission
All users are required to fill the Service Request Form and email it to us at firstname.lastname@example.org. Samples will not be accepted without the form. Samples will be entered into the sequencing queue upon receiving the form and samples.
Full service – Library Preparation and Sequencing
Currently, we only offer library preparation service for genomic DNA sequencing, RNA sequencing, and PCR amplicon sequencing. Users only need to provide us genomic DNA, total RNA, or PCR amplicons, we will perform sample QC, prepare libraries, pool libraries, and perform sequencing runs.
For other sequencing applications, users need to prepare and pool the libraries and bring them to us for sequencing.
Genomic DNA sample submission
For each sample, a minimum of 200 ng genomic DNA is required. The gDNA should be column purified and intact. An agarose gel picture is required and the A260:280 ratio needs to be above 1.8.
RNA-Seq sample submission
For standard RNA-Seq protocols, a minimum of 200 ng total RNA is required. RNA should be column purified and intact (RIN > 8.0; A260:280 ratio > 1.8). We also perform RNA-Seq library preparation with very small input material, as little as 1 ng of total RNA.
PCR amplicon sample submission
For each sample, a minimum of 100 ng PCR amplicons is required. Amplicons should be column purified and the size range should be between 150 bp and 600 bp.
We perform sequencing on user-prepared libraries. Users are responsible to make sure that libraries are compatible with Illumina sequencers. We can also use custom sequencing primers, and again users are responsible for the compatibility with Illumina sequencers.
Pre-made library submission
Users should pool libraries before submitting to us. For each pool, at least 10ul of 10nM material is required. We will check library quality and quantity on TapeStation and Qubit before sequencing.
With user-prepared libraries, we can’t guarantee sequencing quality. If libraries need to be resequenced to obtain enough reads, 50% fees will apply for repeating.
Data Analysis and Delivery
We perform initial data analysis and demultiplexing to generate individual FASTQ file for each library in a pool. We deliver FASTQ files to users through Rockefeller CFS (Central File Storage), or directly to a laboratory’s server if available.
Sequencing Data Retention
Due to the large size and high volume of sequencing data, we have very limited capacity for data retention.
Image files: Illumina sequencers no longer retain image files. Images are analyzed in realtime and immediately deleted by the sequencer software. Users therefore do not have the option of requesting image files.
Intermediate analysis files: The standard procedure is to retain the intermediate files at the GRC only during the troubleshooting phase, which is generally less than a month. If users would like a copy of the intermediate analysis files, they must request this when submitting samples AND provide a portable hard drive for the file transfer.
Analyzed data (FASTQ files): This data are delivered to users’ CFS folders or designated servers, and will be stored at the GRC for six months. Once delivered, it’s users’ responsibility to keep the data files.
If you would like to keep the samples or libraries that have already been sequenced, you need to get them back from us and store in your own freezers. We will keep them in our freezers for one month from the time the sequencing data is delivered. After one month, leftover samples and libraries will be discarded.