# Custom sequencing primers on the MiSeq

For reference, see the [MiSeq System Custom Primers Guide](https://support.illumina.com/downloads/miseq-system-custom-primers-guide-15041638.html).

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.

**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 Guide](https://support.illumina.com/downloads/indexed-sequencing-overview-15057455.html)).

  ![](https://761066130-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FGM9W2DuBTgEXv1ClCm8H%2Fuploads%2Fgit-blob-82e09160149720343660f1777d1b7605afc606aa%2Fimage1.png?alt=media\&token=a33b7e22-304a-488b-9d62-198a8371e44f)

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**.

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