Instructions of sample sheet for demultiplexing

A sample sheet is used for demultiplexing Illumina sequencing data. It contains information about the identity of each sample and the index sequences used to barcode each sample during library preparation. Demultiplexing software reads the index sequences from the sequencing data and assigns each read to the appropriate sample based on the index sequences specified in the sample sheet. This allows researchers to accurately assign sequencing data to specific samples, which is critical for downstream analysis. The sample sheet may also contain other metadata such as information about the sequencing platform, the flow cell used for sequencing, and adapter sequences used during library preparation.

Please download this example file for basic usage: Download

  • Please use comma (,) as separators instead of colon (;).
  • Please be very careful if you enter the sequences of indices manually.
  • Please remove any spaces and special characters (#, +, -, %… etc) in the files. Underline (_) is always preferred.
  • Please refer to your index kit manual and determine whether the sequences should be forward or reverse complements according to the sequencer used.

For any advanced usage of this sample sheet, please refer to the documentation of bcl2fastq 2.0 or ask us.


Sequencing instrumentForward strandReverse complement
iSeq® 100X
1.0 reagentsX
1.5 reagentsX
Rapid ReagentsX
Standard ReagentsX
HiSeq 2500 or 2000
Paired End FlowcellX
Single Read FlowcellX
HiSeq 3000 or 4000
Paired End FlowcellX
Single Read FlowcellX