Home › Cell Biology › Monarch PCR & DNA Cleanup Kit Protocol
Steps
  1. 1 Prepare wash buffer with ethanol 00:09
  2. 2 Prepare DNA sample and add binding buffer 00:25
  3. 3 Load sample and bind DNA to column 01:14
  4. 4 Wash column with buffer twice 01:29
  5. 5 Elute purified DNA from column 01:50
  6. 6 Collect purified DNA in tube 02:14
Cell Biology New England Biolabs

Monarch PCR & DNA Cleanup Kit Protocol

Protocol
Difficulty
intermediate

Steps

1
Prepare wash buffer with ethanol

Add 4 volumes of ethanol to one volume of DNA Wash Buffer according to the bottle label instructions before beginning the protocol.

▶ 00:09
2
Prepare DNA sample and add binding buffer

Start with 20-100 μl of DNA sample and add the recommended amount of DNA Cleanup Binding Buffer according to the protocol card table. Adjust sample volume to at least 20 μl with TE buffer if needed, then mix well by pipetting.

▶ 00:25
3
Load sample and bind DNA to column

Load the sample mixture onto a column seated in a collection tube and spin for 1 minute at 16,000 x g. Remove the column from the collection tube and discard the flow-through.

▶ 01:14
4
Wash column with buffer twice

Re-insert the column into a fresh collection tube and add 200 μl of DNA Wash Buffer, then spin for 1 minute. Repeat the wash step a second time with 200 μl of fresh buffer and spin again for 1 minute.

▶ 01:29
5
Elute purified DNA from column

Transfer the column to a clean microfuge tube without contacting the flow-through. Add 6 or more μl of DNA Elution Buffer to the column matrix center and incubate for 1 minute to maximize yield.

▶ 01:50
6
Collect purified DNA in tube

Spin the column for 1 minute to collect the flow-through, which contains your purified DNA.

▶ 02:14

🚨 Failure Case Library (4) + Submit your own case

critical
No DNA Recovered from Gel Extraction or PCR Cleanup
No DNA is recovered after gel extraction or PCR cleanup procedures. Elution yields no measurable nucleic acid.
💡 3 · ✓ 3
severe
Low Gel Extraction Yield Due to Improper Gel Dissolution
DNA yield from gel extraction is poor. Undissolved agarose particles may be visible or column flow is impeded.
💡 4 · ✓ 4
moderate
Low DNA Yield from Gel Extraction or PCR Cleanup Due to Incomplete Elution
DNA recovery from gel extraction or PCR cleanup is lower than expected based on input amount. Some DNA likely remains bound to column.
💡 4 · ✓ 4
moderate
Low DNA Performance in Downstream Applications Due to Contaminants
Purified DNA fails or performs poorly in downstream applications (transformation, restriction digestion, PCR, sequencing) despite acceptable concentration and A260/280 ratio.
💡 6 · ✓ 6
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