3.2 Load GEM-X Chip

   

  • After removing chip from the sealed bag, use in ≤24 h.

  • Open the lid (gasket attached) of the assembled chip and lay flat for loading.

  • Ensure that the Gel Beads are properly thawed and ready to use.

  • When loading the chip, raising and depressing the pipette plunger should each take ~5 sec. When dispensing, raise the pipette tips at the same rate as the liquid is rising, keeping the tips slightly submerged.

     

    Color Legend

    2: Gel beads
    1: Sample
    3: Oil

     




    The Chromium X/iX (X Series) Chip Holder, Chip Gasket X/iX, and GEM-X chip images shown below are representative and do not show the specific color & label. Chip holder and gasket should be black and blue in color, respectively. Refer to Assemble GEM-X Chip for details.

GEM-X MO Chip, gasket attached

Representative chip image

Pipette technique

 

a. If loading less than 8 samples/chip, add 50% glycerol solution to each unused well in row 1, 2, and 3

  • 60 μl in each unused well in row labeled 1

  • 60 μl in each unused well in row labeled 2

  • 250 μl in each unused well in row labeled 3

 

DO NOT add 50% glycerol solution to the wells in top row labeled NO FILL.

DO NOT use any substitute for 50% glycerol solution.

Glycerol
in GEM-X MO Chip

 

b. Prepare Gel Beads

  • Snap the tube strip holder with the Gel Bead strip into a 10x Vortex Adapter. Vortex 30 sec.

  • Centrifuge the Gel Bead strip for ~5 sec.

  • Confirm there are no bubbles at the bottom of the tubes & the liquid levels are even.

  • Place the Gel Bead strip back in the holder. Secure the holder lid.

Prep Gel Beads


c. Prepare Master Mix + Transposed Nuclei

  • Individual Transposition (nuclei not counted after Post Transposition Wash): 

    • Gently pipette mix 25 μl transposed nuclei in the tube strip and add 40 μl Master Mix to each tube for a total of 65 μl, gently pipette mix 10x, and immediately proceed to step d.

  • Scalable Transposition (nuclei counted after Post Transposition Wash):

    • Add 40 μl Master Mix to each tube of a tube strip.

    • Refer to the Transposed Nuclei Suspension Volume Calculator Table for GEM Generation and add appropriate volume of Transposition Wash Buffer to the Master Mix, gently pipette mix 10x, and centrifuge briefly. DO NOT add Transposition Wash Buffer directly to the nuclei suspension.

    • Add appropriate volume of transposed nuclei to the Master Mix. Gently pipette mix the transposed nuclei before adding.Total volume 65 μl in each tube.

      Gently pipette mix 10x and immediately proceed to step d.


    Any remaining Transposed Nuclei may be maintained on ice for up to 30 min if additional rounds of GEM Generation are required.

 

 

d. Load Row Labeled 1

  • Gently pipette mix the Master Mix + Transposed Nuclei.

  • Using the same pipette tip, dispense 60 μl Master Mix + Transposed Nuclei into the bottom center of each well in row labeled 1 without introducing bubbles.

  • Wait 30 sec.

60 μl Master Mix + Cell Suspension

in GEM-X MO Chip

 

 

e. Load Row Labeled 2

  • Puncture the foil seal of the Gel Bead tubes.

  • Slowly aspirate 60 μl Gel Beads.

  • Dispense into the bottom center of each well in row labeled 2 without introducing bubbles.

  • Wait 30 sec.

60 μl Gel Beads
in GEM-X MO Chip

 

 

f. Load Row Labeled 3

  • Dispense 250 μl Partitioning Oil B into the wells in row labeled 3 by pipetting two aliquots of 125 μl from a reagent reservoir.

Failure to add Partitioning Oil B to the row labeled 3 will prevent GEM generation and can damage the instrument.

250 μl Partitioning Oil B
in GEM-X MO Chip

 

 

g. Prepare for Run

  • Close the lid (gasket already attached). DO NOT touch the smooth side of the gasket. DO NOT press down on the top of the gasket. Keep horizontal to avoid wetting the gasket.

  • Keep the chip horizontal and be careful when moving/setting down the chip to avoid wetting the gasket with oil or spilling oil over the outside of the wells.*


Run the chip in Chromium X series instrument (X/iX) immediately after loading the Partitioning Oil B.

GEM-X MO Chip, closed