DNA/RNA co-extraction from coral tissue (Zymo Quick-DNA/RNA Miniprep Plus)

DNA/RNA extraction
Porites astreoides
Extract DNA and RNA from the same small piece of snap-frozen coral (adult fragment or larvae) using bead-beating lysis and Zymo’s paired DNA and RNA spin columns.
Author

Kevin H. Wong

Published

October 5, 2026

Version 1.0 · last updated 2026-10-05 · Used in 30 notebook entries ↓

At a glance

Input Snap-frozen adult fragment (small clipping of tissue + skeleton) or one vial of larvae (20 larvae), stored at −80 °C
Output ~100 µL genomic DNA (in 10 mM Tris) and ~100 µL total RNA (in water) from the same sample
Kit Zymo Quick-DNA/RNA Miniprep Plus (D7003)
Time ~2 h for 6–10 samples, plus QC (estimate; TODO: confirm)
Tested on Porites astreoides adults and larvae; geoduck tissue (one run)

The sample is lysed in DNA/RNA Shield by bead beating and digested with Proteinase K. It is then passed over a yellow column that binds DNA. The flow-through, which contains the RNA, goes onto a green column with an on-column DNase step. Both columns use the same wash buffers, so the DNA and RNA preps can run side by side.

Materials

Kits and reagents

  • Zymo Quick-DNA/RNA Miniprep Plus kit (D7003): DNA/RNA Shield, Proteinase K + PK Digestion Buffer, DNA/RNA Lysis Buffer, DNA/RNA Prep Buffer, DNA/RNA Wash Buffer, DNase I + DNA Digestion Buffer, yellow (DNA) and green (RNA) spin columns. Kit manual (PDF)
  • 100% ethanol (molecular grade)
  • 10 mM Tris-HCl, pH 8.0, made with RNase-free water (DNA elution)
  • RNase-free water (RNA elution)
  • RNase Away
  • For sterilizing clippers: 10% bleach, DI water, 70–90% ethanol, RNase-free water

Equipment and consumables

  • Bead tubes: 2 mL tubes with 0.5 mm glass beads (Fisher 15340152), or ~0.25 mL loose 0.5 mm glass beads per tube
  • Vortexer with a tube adapter (max speed)
  • Microcentrifuge capable of 16,000 × g
  • Heat block or Thermomixer at 70 °C (pre-warming elution liquids)
  • Sterile clippers, dry ice, 1.5 mL tubes, 0.5 mL PCR tubes for QC aliquots
  • A designated RNase-free bench area

Solutions to prepare (first use of a kit)

Reagent Preparation Storage
DNA/RNA Wash Buffer If supplied as concentrate, add 96 mL 100% ethanol (or 104 mL 95%) to the 24 mL concentrate. The D7003 kit wash buffer is usually ready to use; check the label. Room temp
DNase I Reconstitute the lyophilized enzyme as indicated on the vial; mix by inversion, don’t vortex. Aliquot, −20 °C
Proteinase K 20 mg vial + 1,040 µL PK Storage Buffer (or 5 mg + 260 µL); vortex to dissolve. −20 °C

Before you start

  • Wipe the bench, pipettes and racks with RNase Away. Keep the spray bottle and Kimwipes handy.
  • Pre-warm the 10 mM Tris-HCl and RNase-free water to 70 °C.
  • Thaw the DNase I aliquot on ice (you’ll need it about 30 min in).
  • Label for each sample: one bead tube, one 1.5 mL “lysate” tube, one 1.5 mL “RNA flow-through” tube, two elution tubes (DNA, RNA), and two QC tubes.
  • Keep samples on dry ice until the moment you add Shield.
  • Sterilize the clippers between samples: 10% bleach → DI water → ethanol → RNase-free water.

Procedure

A. Lysis and bead beating

  1. Add ~0.25 mL of 0.5 mm glass beads and 500 µL DNA/RNA Shield to each bead tube.
  2. Add the sample:
    • Adult: clip a small piece of tissue and skeleton (TODO: confirm the size you aim for) straight from dry ice into the tube.
    • Larvae: add the whole vial (20 larvae) to the tube.
  3. Vortex at maximum speed: 1 min for adult fragments, 30 s for larvae.
  4. Transfer 400 µL of the supernatant to a labeled 1.5 mL tube. Avoid mucus clumps and skeleton.
Tip⏸ Safe stopping point

The leftover bead-tube material can go back to −80 °C as a backup. Samples in DNA/RNA Shield are stable, but continue the same day if you can.

B. Proteinase K digestion

  1. Add 40 µL PK Digestion Buffer and 20 µL Proteinase K to the 400 µL lysate. (Ratios: sample : digestion buffer = 10 : 1; digestion buffer : PK = 2 : 1. Scale if you use a different volume.)
  2. Vortex briefly and spin down.
  3. Add an equal volume (460 µL) of DNA/RNA Lysis Buffer. Mix by flicking or brief vortexing.

C. DNA (yellow column)

  1. Load 700 µL of lysate onto the yellow DNA column. Centrifuge 16,000 × g, 30 s.
  2. Keep the flow-through. Transfer it to the labeled “RNA” tube. It contains the RNA (go to part D).
  3. Load the rest of the lysate onto the same yellow column, spin again, and add that flow-through to the RNA tube as well.
  4. Add 400 µL DNA/RNA Prep Buffer. Spin 30 s. Discard the flow-through.
  5. Add 700 µL DNA/RNA Wash Buffer. Spin 30 s. Discard the flow-through.
  6. Add 400 µL DNA/RNA Wash Buffer. Spin 2 min to dry the membrane completely.
  7. Move the column to a labeled 1.5 mL elution tube.
  8. Drip 50 µL warm 10 mM Tris-HCl slowly onto the center of the membrane. Wait 5 min, then spin 30 s.
  9. Add a second 50 µL warm Tris-HCl. Wait 15 min, then spin 30 s. Discard the column (final volume ~100 µL).

D. RNA (green column)

You can start this right after step 9, while the DNA columns are washing.

  1. Add an equal volume of 100% ethanol to the RNA flow-through (≈ 920 µL for a 400 µL starting lysate). Vortex briefly and spin down.
  2. Load 700 µL onto the green RNA column. Spin 30 s and discard the flow-through. Repeat until all the liquid has passed through.
  3. Add 400 µL DNA/RNA Wash Buffer. Spin 30 s. Discard the flow-through.
  4. Make the DNase I master mix: 75 µL DNA Digestion Buffer + 5 µL DNase I per sample, plus one extra.
  5. Add 80 µL of the master mix directly onto the membrane. Incubate 15 min at room temperature.
  6. Add 400 µL DNA/RNA Prep Buffer. Spin 30 s. Discard the flow-through.
  7. Add 700 µL DNA/RNA Wash Buffer. Spin 30 s. Discard the flow-through.
  8. Add 400 µL DNA/RNA Wash Buffer. Spin 2 min.
  9. Move the column to a labeled RNase-free 1.5 mL tube.
  10. Drip 50 µL warm RNase-free water onto the membrane, wait 5 min, and spin 30 s. Repeat once (final volume ~100 µL).

E. Aliquot and store

  1. Take QC aliquots: 10 µL DNA (Qubit + gel) and 5 µL RNA (Qubit + TapeStation) into 0.5 mL tubes.
  2. Store DNA at −20 °C (4 °C if you’re running QC within a day) and RNA at −80 °C.

Clean-up

  • Collect all Zymo buffer waste and tissue in the labeled Zymo extraction waste container. The lysis buffer contains guanidine salts, so don’t mix it with bleach.
  • Wipe the RNase-free area with RNase Away and restock tips.

QC and expected results

Check Method
DNA and RNA concentration Qubit Broad Range DNA and RNA assays (1 µL sample)
DNA integrity 1% agarose gel: expect a high-molecular-weight band with little smear
RNA integrity TapeStation RNA ScreenTape (RIN)

Typical yields from the notebook (100 µL elutions):

Sample type DNA (ng/µL) RNA (ng/µL) Source
P. astreoides adult fragment, bead beating 7–23 22–38 2020-10-22, n = 6
P. astreoides larvae (20 per vial), bead beating 14–48 34–57 2020-08-12, n = 6
P. astreoides larvae, PK digestion only (no beads) 19–56 24–75 2020-08-06, n = 10

Troubleshooting

Problem Likely cause Fix
RNA degraded (low RIN) but DNA fine Sample was an airbrushed homogenate; RNA from homogenates came out too degraded in the first test. Use airbrushed homogenates for DNA only. For RNA, bead-beat a clipping taken straight from −80 °C (2019-02-13).
Hard to pipette the supernatant; column clogs Mucus clumps or skeleton fragments Spin the bead tube briefly (8,000 × g, 30 s) and take the supernatant from the top (2020-08-06).
Very dilute lysate, low yield from fragments Too much Shield for a small clipping (1,000 µL was too much) Use 500 µL Shield per bead tube.
DNA contamination in RNA DNase step skipped or too short Use the full 15 min on-column DNase with fresh master mix.
Low DNA yield Membrane not fully dry, or elution not warm Do the 2 min dry spin. Use Tris pre-warmed to 70 °C and the 15 min second incubation.

Notes by sample type

  • Larvae. Bead beating (30 s) replaced the original 1.5 h Proteinase K digestion at 55 °C (1,000 µL Shield + 100 µL PK buffer + 50 µL PK). Bead beating is faster and gave similar yields.
  • Adult fragments. Bead-beat for 1 min. Airbrushed homogenates work for DNA but not RNA (see Troubleshooting).
  • Other species. One run included geoduck tissue alongside Porites (2020-07-30).

References

Notebook entries using this protocol

Date Title Project
2020-12-03 20201202 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-30 20201126 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-25 20201125 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-23 20201119 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-23 20201120 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-18 20201117 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-12 20201110 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-12 20201111 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-11-09 20201106 DNA RNA Extractions Porites July Bleaching Experiment and WGBS samples Porites July Bleaching
2020-10-29 20201029 DNA RNA Extractions Porites July Bleaching Experiment and WGBS samples Porites July Bleaching
2020-10-27 20201027 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-10-26 20201025 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-10-23 20201022 DNA RNA Extractions Porites July Bleaching Experiment Porites July Bleaching
2020-08-20 DNA RNA Extractions on P. astreoides larvae - BEAD BEATING Batch 5 Porites Thermal Transplant
2020-08-19 DNA RNA Extractions on P. astreoides larvae - BEAD BEATING Batch 4 Porites Thermal Transplant
2020-08-16 DNA RNA Extractions on P. astreoides larvae - BEAD BEATING Batch 3 Porites Thermal Transplant
2020-08-13 DNA RNA Extractions on P. astreoides larvae - BEAD BEATING Batch 2 Porites Thermal Transplant
2020-08-12 DNA RNA Extractions on P. astreoides larvae - BEAD BEATING Porites Thermal Transplant
2020-08-06 DNA-RNA extractions Porites Thermal Transplant Larvae BATCH 1 Porites Thermal Transplant
2020-08-05 DNA-RNA Extractions Porites astreoides homogenates, larvae, and adult fragments Porites Thermal Transplant
2020-07-31 DNA-RNA-Extraxtion-Thermal-Transplant-Porites-Homogenates Porites Thermal Transplant
2020-07-30 DNA-RNA-Extractions-Thermal-Transplant-Porites-and-Geoduck Porites Thermal Transplant
2020-01-13 DNA RNA Extractions Thermal Transplant ADULT AND LARVAE Trial 4 Porites Thermal Transplant
2019-12-05 DNA RNA Extractions Thermal Transplant Porites LARVAE Trial 1 Porites Thermal Transplant
2019-12-05 DNA RNA Extractions Thermal Transplant Porites Trial 3 Porites Thermal Transplant
2019-12-04 DNA RNA Extractions Thermal Transplant Porites Trial 1 Porites Thermal Transplant
2019-12-04 DNA RNA Extractions Thermal Transplant Porites Trial 2 Porites Thermal Transplant
2019-12-04 DNA RNA Extractions Thermal Transplant Porites Trial 2 Porites Thermal Transplant
2019-03-13 Zymo-DNA-RNA-Extract-P.astreoides-Genome Porites astreoides Genome
2019-02-13 Zymo DNA RNA Extraction Protocol  
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Changelog

Date Version Change
2026-10-05 1.0 Written up as a standalone protocol from the notebook entries below
2020-10 – Bead-beating lysis applied to adult fragments (Porites July Bleaching)
2020-08 – Bead beating replaced the 1.5 h Proteinase K digestion for larvae
2019-12 – Shield + glass-bead “soft homogenization” and the paired DNA/RNA column workflow (Thermal Transplant trials)
2019-02 – First test of the kit on airbrushed homogenates vs. fragments
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