The Illumina NextSeq 550 System offers several precise read lengths, also known as "read sizes," to accommodate a variety of sequencing applications. These read lengths are defined by the number of base pairs (bp) in each sequence read.
The primary read lengths available for the NextSeq 550 System include:
- 1 × 75 bp (single-end reads): A single read of 75 base pairs.
- 2 × 75 bp (paired-end reads): Two reads, each 75 base pairs long, originating from opposite ends of a DNA fragment.
- 2 × 150 bp (paired-end reads): Two reads, each 150 base pairs long, from opposite ends of a DNA fragment.
Understanding NextSeq Read Lengths and Output
The choice of read length directly impacts the total data output (in Gigabases, Gb) per flow cell, as well as the suitability for different experimental designs. The NextSeq 550 System utilizes specific kits, Mid-Output and High-Output, which determine the overall data capacity for a given read length.
Here's a breakdown of the output per flow cell for various read lengths, illustrating how read size relates to data generation:
| Read Length | NextSeq 550 System Mid-Output Kit | NextSeq 550 System High-Output Kit |
|---|---|---|
| 1 × 75 bp | N/A | 25–30 Gb |
| 2 × 75 bp | 16.25–19.5 Gb | 50–60 Gb |
| 2 × 150 bp | 32.5–39 Gb | 100–120 Gb |
Key aspects of read length options:
- Single-End (1x): In a 1 × 75 bp run, the sequencer reads a single strand of DNA for 75 base pairs. This is often sufficient for applications like gene expression profiling (RNA-Seq) or genotyping where directional information from both ends of a fragment is not critical.
- Paired-End (2x): Paired-end sequencing involves reading both ends of a DNA fragment. For instance, a 2 × 75 bp run means the system sequences 75 bp from one end and then 75 bp from the other end of the same DNA molecule. Similarly, 2 × 150 bp provides 150 bp from each end. Paired-end reads are highly valuable for:
- De novo assembly: Helps bridge gaps and resolve repetitive regions.
- Variant detection: Improves accuracy by providing two independent reads covering a specific region.
- Gene fusion detection: Can span breakpoints in chromosomal rearrangements.
- RNA-Seq: Provides better alignment and quantification for transcripts, especially for alternative splicing analysis.
The 2 × 150 bp read length offers the highest data output and is generally preferred for applications requiring deeper coverage or more comprehensive genomic information, such as whole-genome sequencing or exome sequencing.
