NextSeq 1000/2000 FAQs

The options are based on the available DRAGEN pipelines:

• [DRAGENEnrichment_Settings] and [DRAGENEnrichment_data]
• [DRAGENGermline_Settings] and [DRAGENGermline_data]
• [DRAGENRNA_Settings] and [DRAGENRNA_data]
• [DragenSingleCellRNA_Settings] and [DragenSingleCellRNA_data]

• Cloud mode—Plan your run with Instrument Run Setup on BaseSpace Sequence Hub. The run is selected from a list of planned runs in NextSeq 1000/2000 Control Software. During sequencing, cBCL data are uploaded to BaseSpace Sequence Hub. The selected analysis workflow is initiated automatically within the cloud. Run data and analysis results are also provided in the cloud.
• Hybrid mode—Plan your run with Instrument Run Setup on BaseSpace Sequence Hub. The run is selected from a list of planned runs in NextSeq 1000/2000 Control Software. The selected analysis workflow is then initiated through the on-instrument DRAGEN.
• Local mode—Plan your run with a sample sheet v2 file format locally. The selected analysis workflow is initiated automatically through the on-instrument DRAGEN or manually through BaseSpace Sequence Hub apps after run completion when Proactive, Run Monitoring and Storage is selected.
• Standalone mode—Plan your run without a sample sheet, following the instructions in NextSeq 1000/2000 Control Software to generate cBCL data.

Analysis is available through the on-board DRAGEN Bio-IT Platform or on the cloud using BaseSpace Sequence Hub. The following DRAGEN pipelines are available:

• DRAGEN BCL Convert
• DRAGEN Enrichment
• DRAGEN Germline
• DRAGEN RNA
• DRAGEN Single Cell RNA

For each read, entering a minimum of 26 cycles and a maximum of 151 cycles helps ensure data quality. The exact number of cycles depends on your experiment. NextSeq 1000/2000 Control Software requires at least 1 cycle for Read 1, but displays a warning when the number of cycles in Read 1 is less than 26. The total number of cycles for Read 1, Index 1, Index 2, and Read 2 cannot be greater than the number of cycles supported by the kit plus 27. NextSeq 1000/2000 Control Software will display a warning when Index 1 and Index 2 are less than 6 cycles. The warning will not be shown if Index 1 or Index 2 is 0 cycles.

Run duration depends on the type of reagents and number of cycles. For example run times, see Specifications for the NextSeq 1000 and NextSeq 2000 Systems.

The NextSeq 1000/2000 P2 Reagents can generate up to 120 Gb per sequencing run and the NextSeq 2000 P3 Reagents can generate up to 330 Gb per sequencing run. For detailed performance parameters, see Specifications for the NextSeq 1000 and NextSeq 2000 Systems.

Depending on the sequencing mode selected, sample information is entered using Instrument Run Setup BaseSpace Sequence Hub or a Sample Sheet v2 template (see Product Files).    

The following applications are targeted for the NextSeq 1000/2000 system:

• Small whole-genome sequencing
• Whole-exome sequencing
• Single-cell RNA-Seq

The loading volume is 20 µl and the loading concentration varies by library type:

For other library types, 650 pM is the recommended starting loading concentration. Optimize this concentration over subsequent runs to identify a loading concentration that consistently yields data that meets specifications.

The number of libraries you can sequence at one time depends on the desired coverage. Using supported Illumina kits, you can multiplex up to 384 libraries. For coverage considerations, see the Sequencing Coverage Calculator.

Custom primers are supported for the NextSeq 1000/2000 P2 Reagents kit and the NextSeq 1000/2000 P3 Reagents kit. If using the NextSeq 1000/2000 P2 (v2) Reagents kit (catalog # 20043738, 20043737, and 20043736), you will need to use a custom recipe to use custom primers. See Product Files for more information. 

Yes, NextSeq 1000/2000 P2 Reagents and NextSeq 1000/2000 P3 Reagents include all consumables to support both single-read and paired-end sequencing. During run setup, you can configure the run to generate either single-read or paired-end data.