Prepare Dry Cartridge

MiSeq i100 Plus consumables are shipped and stored at room temperature. Thawing is not required. Before loading the libraries into the dry cartridge, dilute libraries and optionally spike in PhiX. Libraries are denatured automatically onboard the instrument.

Always perform a quality control analysis and optimize the loading concentration for your library.

Dilute Libraries

1.

Use scissors to cut open the wet cartridge foil packaging to retrieve the RSB and KLD tubes. Set the tubes aside.

2.

Dilute libraries to 10x loading concentration using Resuspension Buffer (RSB).

Example: For a final loading concentration of 100 pM, dilute to 1 nM.

3.

Vortex at the highest setting for 3 seconds, and then centrifuge briefly.

4.

[Optional] PhiX spike in as follows.

a.

Dilute PhiX to 6x library loading concentration with RSB.

Example: For a final loading concentration of 100 pM, dilute PhiX to 0.6 nM.

b.

Add 1uL of this dilution to the 10x loading concentration library mixture.

The volumes produce approximately a 2% PhiX spike in. The percentage varies depending on the library quality and quantity.

6.

Vortex at the highest setting for 3 seconds, and then centrifuge briefly.

7.

Store mixture on ice until ready for use.

The diluted library solution is stable up to 6 hours when stored on ice or at 4ºC.

Load Libraries

1.

Put on a new pair of powder-free gloves to avoid contamination.

2.

Use scissors to cut open the dry cartridge foil packaging.

Use the dry cartridge within 4 hours of opening the foil packaging.

3.

Remove the dry cartridge from the package.

Grasp the dry cartridge by the sides to avoid touching the flow cell.

4.

Discard the foil package appropriately.

5.

Using a clean pipette tip, pierce the foil seal covering the reagent well labeled Library.

6.

Pipette 250 µl diluted library solution into the Library well in the dry cartridge.

7.

Optional Pipette custom primer into the appropriate port on the dry cartridge. Refer to Custom Primers.