Home Resources Reagent & Buffer Preparation Guide for Monarch Spin RNA Isolation Kit (Mini) (NEB #T2110)

Reagent & Buffer Preparation Guide for Monarch Spin RNA Isolation Kit (Mini) (NEB #T2110)

  • Always keep columns tightly sealed in the provided bag.
  • Always keep all buffer bottles tightly closed when not in use.
  • Store kit components at the recommended storage conditions on component labels and in the product manual

 

Reagents Supplied by the User


Equipment

  • Benchtop microcentrifuge
  • Water bath, heat block or thermomixer
  • Bead homogenizer
  • Vacuum manifold (for the vacuum manifold protocol)
  • Vacuum pump (for the vacuum manifold protocol)

 

Reagents/supplies

  • RNase-free 1.5 ml microfuge tubes
  • Ethanol (≥ 95%), Molecular Biology Grade
  • Isopropanol (100%) for whole blood samples
  • Glycanases for microbes, for example, NEBExpress T4 Lysozyme (NEB #P8115)
  • Xylene or similar for deparaffinization of FFPE tissue.

 

Reagent Preparation

Rehydration of Monarch DNase I, Lyophilized

Monarch DNase I is a proprietary lyophilized mix that is stable at room temperature in the supplied dry cake format. For rehydration and subsequent storage guidance, follow the instructions below:

  1. Place the glass vial containing lyophilized enzyme on the RNase-free benchtop.

  2. Unscrew the cap and place the cap upside down on the benchtop.

  3. Remove the grey rubber stopper and place it securely inside the inverted screw cap.

  4. Using a pipette, dispense the appropriate volume of the supplied Nuclease-free Water onto the lyophilized cake.

    1. For T2104-21, dispense 220 µl Nuclease-free Water

    2. For T2104-1, dispense 550 µl Nuclease-free Water

  5. Pipette mix 8-10 times adjusting the tip positioning inside the vial such that all the lyophilized material gets hydrated. Do not vortex.

  6. Place the vial back on the bench.

  7. The solution will turn clear within 15 seconds. Let it stay for an additional minute.

  8. The rehydrated DNase I is ready for use in the RNA isolation protocol.

  9. Store the rehydrated DNase I at -20°C. Avoid more than three freeze-thaw cycles. We strongly recommend storing the rehydrated DNase I in appropriately sized aliquots based on the user’s processing needs.

 

Preparation of Monarch Buffer BX

Monarch Buffer BX serves as the RNA priming reagent and is provided as a concentrate. Before using in the workflow, follow the steps below for reconstitution in the sequence listed:

  1. Add the appropriate volume of the supplied Nuclease-free Water to the reagent bottle
    1. To T2041-11, add 2 ml Nuclease-free Water

    2. To T2041-22, add 10 ml Nuclease-free Water

  2. Add the appropriate volume of ≥ 95% ethanol (user-supplied)

    1. To T2041-11, add 2 ml ethanol

    2. To T2041-22, add 10 ml of ethanol

 

Preparation of Monarch Buffer WZ

Monarch Buffer WZ serves as an RNA wash buffer and is provided as a concentrate. Before using:

  1. Add the appropriate volume of ≥ 95% ethanol (user-supplied)

    1. To T1115-1, add 20 ml ethanol

    2. To T1115-3, add 104 ml ethanol