General Questions
Please see the services page for a detailed list of projects we support. If your project design is not listed, please contact ccrsfhelp@mail.nih.gov
or the Sequencing Facility director, to discuss the feasibility of a custom project.
All NIH research labs are eligible to order services through the Sequencing Facility. Labs outside of CCR and NIAID will have overhead charges added.
Please complete a sequencing proposal form at NAS request. There will be an option for Illumina, which you should select for all short read and single-cell projects, and Long Read, which you should select for all PacBio, ONT and Bionano projects. You may also contact ccrsfhelp@mail.nih.gov to discuss the available platforms and best choice for your project.
Before sequencing, we will perform an internal QC to confirm the information in the sample manifest and notify you if any samples do not meet minimum sequencing requirements. You will then be able to choose whether to resubmit those samples or continue and sequence them at your own risks. You will be notified again when the analysis on each sample is completed and available for download.
Illumina Questions
Before submitting samples, ensure that the sequencing project has been discussed with the Sequencing Facility team and the NAS request submitted. Illumina service is listed under Sequencing Facility –Illumina (CCR).
You may then submit samples by delivering them at the ATRF Room D3040 (instructions for sample delivery. Be sure to include a sample manifest form with your submission as well as to send an electronic version of the form to Jyoti Shetty prior to sending your samples.
Sample Quantity/Quality Requirements and Recommendations:
All samples are shipped in dry ice and the individual (1.5-2 ml) tubes are labeled clearly overhead charges added
| Type of library | Minimum DNA/RNA Requirement for Library Construction | Recommended DNA/RNA for Optimal Library Construction | Maximum Sample Volume Requirement for Library Construction | Additional requirements |
| ChIP DNA Sequencing | 5 ng | 10 ng | 30 µL | Bulk of the DNA fragments in the 100-300 bp range |
| gDNA Sequencing | 100 ng | 1 µg | 30 µL | DNA should be as intact as possible with no contamination, OD260/280 1.8–2.0 |
| mRNA Sequencing | 25 ng | 1 µg | 30 µL | RIN should be at least 8.0, DNase treated |
| mRNA ultralow | 100 pg | 10 ng | 10 µL | RIN should be at least 8.0, DNase treated |
| microRNA Sequencing | 100 ng | 1 µg | 6 µL | |
| Total RNA Sequencing | 10 ng | 1 µg | 10 µL | DNase treated, FFPE and degraded RNA can be used; DV200 < 30% not recommended |
You can use any extraction protocol as long as the DNA/RNA samples meet our sample requirements.
Here are the requirements for ATAC seq from frozen cells:
- Bulk ATAC-seq requires each sample to be present as a replicate. Triplicates are better.
- Cells for ATAC-seq should be cryopreserved at high viability. Please ensure that the cells are cryopreserved properly in freezing medium.
- Please send a minimum of 2 million cells per sample.
- Please ship the cells in dry ice.
PacBio Questions
Before submitting samples, ensure that the sequencing project has been discussed with the Sequencing Facility team and the NAS request submitted. PacBio service is listed under Sequencing Facility – Long Read Technology (CCR). You may then submit samples by delivering them at the ATRF Room D3040 (instructions for sample delivery).
Be sure to include a sample manifest form with your submission as well as to send an electronic version of the form to caroline.fromont@nih.gov prior to sending your samples.
All samples must be sent in a 1.5 ml or 2 ml tubes.
Some requirements might be project dependent such as input of DNA if multiplexing or sequencing on multiple SMRTcells. Please contact us for more details.
Quality and quantity requirements are listed in the table below:
| Type of library | Minimum DNA/RNA Requirement for Library Construction | Recommended DNA/RNA for Optimal Library Construction | Maximum Sample Volume Requirement for Library Construction | Quality Requirements |
| WGS | 1.5 µg | 5 µg | n/a | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| Amplicons (< 5000 bp) | 200 ng | 500 ng | n/a | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| Amplicons (> 5000 bp) | 300 ng | 800 ng | n/a | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| HLA (Class I) | 250 ng | 1 µg | 20 ng/µL | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| 16S | 2 ng | 10 ng | 500 pg/µL | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| MAS Single Cell | 15 ng | 50 ng | 1 ng/µL | OD260/280:1.8-2.0 OD260/230:1.7-2.2 |
| WTS | 300 ng | 1 µg | 50 ng/µL | RIN ≥ 8.0 |
Please complete a sequencing proposal form at NAS request. There will be an option for Illumina, which you should select for all short read and single-cell projects, and Long Read, which you should select for all PacBio, ONT and Bionano projects. You may also contact ccrsfhelp@mail.nih.gov to discuss the available platforms and best choice for your project.
Before sequencing, we will perform an internal QC to confirm the information in the sample manifest and notify you if any samples do not meet minimum sequencing requirements. You will then be able to choose whether to resubmit those samples or continue and sequence them at your own risks. You will be notified again when the analysis on each sample is completed and available for download.