HiSeq 1000/1500/2000/2500 rehybridization quick reference chart

Rehybridization of primers to a flow cell allows the current sequencing read to be repeated from the first base of that read. Primer rehybridization might be required to resolve various issues, including fluidic issues that affect intensity for one or more lanes on a flow cell or when incorrect run parameters are entered during run setup.

Priming of the reagent lines might be necessary before rehybridization. The following tables provide requirements for priming before rehybridization and list the paired-end rack reagents required for the rehybridization.

Make sure that the remaining SBS reagents are sufficient to complete the sequencing run. Depending on how many cycles of the read were completed before the run was stopped, SBS reagents might need replenishing.

In addition to the SBS reagents, the rehybridization process requires additional primers and reagents, depending on the type of rehybridization performed. Rehybridization Kit details:

• TruSeq v3: Read 1 rehybridization requires the TruSeq v2 cBot Multi-Primer Re-Hybridization Kit (Catalog No. GD-304-2001). Contact Illumina Technical Support for questions regarding reagents for the other types of TruSeq v3 rehybridizations.

• HiSeq v4 rehybridizations require the HiSeq Multi-Primer Rehybridization Kit v4 (Catalog No. GD-403-4001).

• HiSeq Rapid v2 rehybridizations require the HiSeq Rapid Rehybridization Kit (Catalog No. GD-404-1001).

If a wash was not performed before rehybridization:

If a wash was performed before rehybridization:

\*HP8 and HP7 are not compatible with Nextera libraries. HP12 and HP11 are compatible with all library types.

For more detailed information on performing the various types of rehybridization runs, see:

• Primer Rehybridization on a HiSeq High Output Flow Cell

• Primer Rehybridization on a HiSeq Rapid Flow Cell

For more information on rehybridization runs using the HiSeq X, refer to the Read 1 Primer Rehybridization on a HiSeq X Flow Cell guide.

For information on rehybridization runs using the HiSeq 3000/4000, refer to the Read 1 Primer Rehybridization on a HiSeq 3000/4000 Flow Cell guide.

For any feedback or questions regarding this article (Illumina Knowledge Article #1525), contact Illumina Technical Support [e-mail].