Next Generation Sequencer

A NextSeq 500/550 generates sequencing reads for all samples in the quantified library pool and integrates with the VeriSeq NIPT Solution v2 – RUO via the Onsite Server. Sequencing data are evaluated by the Analysis Handler of the VeriSeq NIPT Assay Software.

Consider the following when integrating a NGS with the VeriSeq NIPT Solution v2 – RUO.

• Data storage integration.
• Analysis throughput capacity.
• Network traffic constraints.

The VeriSeq NIPT Assay Software requires a next generation sequencer capable of generating sequencing data on the prepared library pool according to the following specifications:

• Production of 2 x 36 paired‐end reads.
• Compatible with index adapters in the TG VeriSeq NIPT Sample Prep Kit.
• Two channel chemistry.
• Automatic production of base call (BCL) files.

A typical sequencing run for the VeriSeq NIPT Solution v2 – RUO requires 25–30 GB for NextSeq 500/550 data. The actual data size may vary based on the final cluster density. The Onsite Server provides more than 7.5 TB of storage space, which is enough space for about 300 sequencing runs (7,500 / 25 = 300).

For data storage purposes, map the NextSeq 500/550 to the Onsite Server for one of the following methods:

• Use the Onsite Server as a temporary data repository. In this configuration, the instrument is mapped directly to the server and persists data on the local drive.
• For a high throughput lab, use network-attached storage (NAS). Configure the NextSeq 500/550 to persist the sequencing data directly to a specific location on the NAS.

In this setup, configure the Onsite Server to monitor the specific NAS location that enables the server to monitor upcoming sequencing runs. Multiple NextSeq 500/550s can be added to increase sample throughput. For more information on how to map the server to the NAS, refer to Manage a Shared Network Drive.

For more information on how to map NextSeq 500/550s to the server or to the NAS, refer to the user guide for the system.

The VeriSeq NIPT Analysis Pipeline typically processes data for a single sequencing run in approximately 5 hours. When expanding the lab for sample throughput, consider that a single server is able to process a maximum of four runs per day, which totals to 48 samples x 4 = 192 samples per day. Contact Illumina Technical Support for additional throughput solutions.

The VeriSeq NIPT Solution v2 – RUO uses the lab Local Area Network (LAN) for data throughput between the NextSeq 500/550, Onsite Server, and NAS (if configured). When expanding for sample throughput, consider the following IT infrastructure traffic constraints:

• The average data traffic of approximately 25 GB generated over approximately 10 hours is about 0.7 MB/sec per sequencer.
• The lab infrastructure may also support other sources of traffic that must be factored in.

If you use a NextSeq 500/550 that has the VeriSeq NIPT Local Run Manager module, prepare for sequencing as follows.

1. In VeriSeq NIPT Local Run Manager, select Create Run.
2. From the drop-down menu, select VeriSeq NIPT.
3. Complete the following fields:
• Run Name
• Run Description (optional)
• Pool Barcode

The pool barcode entered in the Local Run Manager module must match the pool barcode entered in the Workflow Manager. Incorrect run configurations are rejected by the VeriSeq NIPT Assay Software and may require resequencing. Pool barcodes must be new and unique. Analysis fails if the barcode is associated with a previously analyzed batch.

4. Select Save Run.

After you finish setting up the run, you can initiate the run using the instrument software.