Chemistry and imaging on NextSeq 500/550
Chemistry and base calling
The NextSeq 500/550 uses two-color chemistry, enabling very fast imaging.
Signal in only the red channel results in a C call, whereas signal in only the green channel results in a T call.
Signal in both channels yields an A call, and a G call results from the absence of signal in either channel.
Because of this, there must be at least one non-G base in the first 5 cycles of sequencing for a cluster to be registered during template generation.
Additionally, a G overcall in a run is generally indicative of a failure of clustering (or paired-end turnaround) or very low clustering.
Reference:
Imaging
Review the section titled 'Flow Cell Tiles' of the NextSeq 500 and NextSeq 550 Sequencing System Guide.
Camera numbering
The NextSeq 500/550 uses six cameras to image the flow cell.
Cameras are numbered 1-6. Cameras 1-3 image lane one.
Cameras 4-6 image lane three.
After lanes 1 and 3 are imaged, the imaging module moves on the X-axis to image lanes 2 and 4.
Lane numbering
Lanes 1 and 3, called lane pair A, are imaged at the same time.
Lanes 2 and 4, called lane pair B, are imaged when imaging of lane pair A is complete.
A Lane Pair A—Lanes 1 and 3.
B Lane Pair B—Lanes 2 and 4.
Swath numbering
Each lane is imaged in three swaths.
Swaths are numbered 1-3 for high output flow cells.
Tile numbering
There are 12 tiles in each swath of each camera segment.
Tiles are numbered 01-12, regardless of swath number or camera segment, and represented in two digits.
The complete tile number includes 5 digits to represent the location, as follows:
Surface—1 represents the top surface; 2 represents the bottom surface.
Swath—1, 2, or 3.
Camera—1, 2, 3, 4, 5, or 6.
Tile—01, 02, 03, 04, 05, 06, 07, 08, 09, 10, 11, or 12.
Example: Tile number 12508 indicates top surface, swath 2, camera 5, and tile 8.
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