Custom sequencing primers on the MiSeq

For reference, see the MiSeq System Custom Primers Guide.

Custom sequencing primer positions:

• Custom Read 1 primer: position 18

• Custom Index Read primer: position 19

• Custom Read 2 primer: position 20

When adding custom primers to the custom primer wells, pipette 600 µl of custom primer at 0.5 µM into the designated well. Use HT1 (Hybridization Buffer) to dilute the custom primer, if needed.

Can only one custom index primer be used?

• Yes, only index 1 may be custom or standard. Index 2 is primed off the oligo flow cell lawn (see the Indexed Sequencing Overview).

Alternatively, custom primers can be spiked into the existing Illumina sequencing primers. For further information, search for Knowledge Base article Spiking custom primers into the Illumina sequencing primers.

For any feedback or questions regarding this article (Illumina Knowledge Article #2414), contact Illumina Technical Support [e-mail].