The Monarch PCR & DNA Cleanup Kit rapidly and reliably purifies up to 5 μg of concentrated, high-quality DNA from PCR and other enzymatic reactions. The kit utilizes a bind/wash/elute workflow with minimal incubation and spin times. The columns ensure zero buffer retention and no carryover of contaminants, enabling elution of sample in volumes as low as 6 μl. The buffers provided have been optimized, and do not require monitoring of pH. Eluted DNA is ready for use in restriction digests, DNA sequencing, ligation and other enzymatic manipulations. Designed with sustainability in mind, Monarch kits use significantly less plastic and responsibly-sourced, recyclable packaging. The protocol can also be modified to enable the purification of smaller DNA fragments, including oligonucleotides and ssDNA.
DNA from PCR reactions can be purified after amplification to remove polymerases, primers, detergents, dNTPs, etc.
Enzymatic reaction cleanup
Restriction enzymes and modifying enzymes such as ligases, kinases, nucleases, phosphatases are efficiently removed, allowing for effective desalting and concentration of the DNA sample.
cDNA cleanup
DNA/RNA complexes can be purified post-reverse transcription/amplification to enable removal of the RT and polymerase as well as nucleotides.
Labeling cleanup
Unincorporated radiolabeled or fluorescently labeled nucleotides can be removed from the DNA substrate
Plasmid cleanup
Plasmid preps from unknown sources may contain inhibitors and unwanted contaminants. Purification and concentration can be easily achieved using this kit.
Monarch DNA Cleanup Column Design Monarch columns are designed for performance Monarch columns are designed without a frit, which eliminates buffer retention and the risk of carryover contamination, providing fast, worry-free DNA purification.
Specifications
DNA Sample Type:
DNA from PCR and other enzymatic reactions (e.g., restriction digests, kinase reactions, ligations).
ssDNA or dsDNA oligonucleotides from enzymatic reactions can also be purified using the Oligonucleotide Cleanup Protocol.
Binding Capacity:
up to 5 μg
DNA Size Range:
~50 bp to 25 kb
DNA ≥ 15 bp to 25 kb (dsDNA) and DNA ≥ 18 nt to 10 kb (ssDNA) can also be purified using the Oligonucleotide Cleanup Protocol.
Typical Recovery:
DNA (50 bp to 10 kb): 70–90%
DNA (11–23 kb): 50–70%
ssDNA ≥ 18 nt and dsDNA ≥ 15 bp: 70-85%
Elution Volume:
≥ 6 μl
Purity:
A260/280 > 1.8 and A260/230 > 1.8
Protocol Time:
5 minutes of spin and incubation time
Compatible Downstream
Applications:
ligation, restriction digestion, labeling and other enzymatic
manipulations, library construction and DNA sequencing.
Monarch PCR & DNA Cleanup Kit (5 µg) Protocol Monarch PCR & DNA Cleanup Kit (5 μg) performs equivalently to the leading supplier Preps were performed according to recommended protocols. 1 μg of a 3 kb DNA fragment was incubated with 1 μM primers and OneTaq® Quick-Load® 2X Master Mix (NEB #M0486). DNA was eluted in 20 μl (NEB) and 40 μl (Qiagen) Elution Buffer. Half of the total elution volume was digested with 5 units of DraIII-HF® (NEB #R3510). The digest and the unused portion of the elution were resolved on a 1% w/v agarose gel along with a representative sample of the starting material. Monarch PCR & DNA Cleanup Kit (5 μg) removes low molecular weight primers from dsDNA samples Three independent amplicons (267 bp, 520 bp, 1003 bp) were spiked with two oligonucleotides (16-mer, 24-mer) to a final concentration of 1 μM. Half of each mix was purified with the Monarch PCR & DNA Cleanup Kit (5 μg) following the included protocol. Equivalent fractions of the original mixture and the eluted material were resolved on a 20% TBE acrylamide gel at 100V for one hour and stained with SYBR Green II.
This product is related to the following categories:
The kit should be stored at room temperature. Always keep buffer bottles tightly closed and keep columns sealed in the enclosed zip-lock bag. For information regarding the composition of buffers, please consult the Safety Data Sheets. Proper laboratory safety practices should be employed, including the use of lab coats, gloves and eye protection.
Reagents added incorrectly. Check protocol to ensure correct buffer reconstitution, order of addition for buffers, and proper handling of column flow-through and eluents.
Incomplete elution during prep. Ensure the DNA Elution Buffer is delivered directly to the center of the column so that the matrix is completely covered and elution is efficient. Larger elution volumes and longer incubation times can increase yield of DNA off the column at the cost of dilution of the sample and increased processing times. For typical fragments below 10 kb, the recommended elution volumes and incubation times should be sufficient, unless the maximal yield is desired. For the purification of larger fragments, heating the DNA Elution Buffer to 50°C prior to eluting and extending the incubation time after buffer addition to 5 minutes can improve yield. Additionally, multiple rounds of elution can be employed to increase the amount of DNA eluted, at the expense of dilution of the sample.
Low DNA Performance
Ethanol has been carried-over. Ensure final wash spin time is 1 minute to ensure complete removal of the wash buffer from the column, and be careful when transferring the column to a new tube for elution step to ensure column tip does not contact column flow-through.
Trace amounts of salts that produce low OD260/230 ratios can also be carried over during the elution step. Be careful when transferring column to new tube for elution step to ensure the column tip does not contact column flow-through.
Monarch PCR & DNA Cleanup Kit protocol
Learn how to isolate DNA from your enzymatic reactions, including PCR, using the Monarch PCR & DNA Cleanup Kit (5 µg).
Tips for using the Monarch PCR & DNA Cleanup Kit
Optimize your DNA isolation from PCR and other enzymatic reactions with our quick tips for using the Monarch PCR & DNA Cleanup Kit.
How to recycle your Monarch Kit components
Learn how you can easily recycle all of the components in your Monarch Kits.
Fomenkov, A., Vincze, T., Mersha, F., Roberts, R.J. (2018) Complete genome sequence and methylome analysis of Bacillus caldolyticus NEB414. Genome Announ.; 6 (6), e01605-17. PubMedID: 29439055 , DOI: 10.1128/genomeA.01605-17
Bornikoel J, Staiger J, Madlung J, Forchhammer K, Maldener I. (2018) LytM factor Alr3353 affects filament morphology and cell-cell communication in the multicellular cyanobacterium Anabaena sp. PCC 7120 Mol Microbiol; PubMedID: 29437253
Wei, H., Yan, B., Gagneur, J., Conradt, B. (2017) Caenorhabditis elegans CES-1 snail represses Pig-1 MELK expression to control asymmetric cell division. Genetics; 206(4), PubMedID: 28652378
James L. Dimond,Sanoosh K. GamblewoodSteven B. Roberts (2017) Genetic and epigenetic insight into morphospecies in a reef coral. Mol Ecol; PubMedID: 28753237
Olga De Castro , Maria Comparone, Antonietta Di Maio, Emanuele Del Guacchio, Bruno Menale, Jacopo Troisi, Francesco Aliberti, Marco Trifuoggi , Marco Guida (2017) What is in your cup of tea? DNA Verity Test to characterize black and green commercial teas. PLOS One; PubMedID: 28542606
Su M, Kirchner A, Stazzoni S, Müller M, Wagner M, Schröder A, Carell T. (2016) 5-Formylcytosine Could Be a Semipermanent Base in Specific Genome Sites. Angew Chem Int Ed Engl; PubMedID: 27561097
Anismrita Lahon, Ravi P. Arya, Alexander R. Kneubehl, Megan B. Vogt, Natalie J. M. Dailey Garnes, and Rebecca Rico-Hesse,* (2016) Characterization of a Zika Virus Isolate from Colombia. PLoS One; PubMedID: 5031432
Publications
Wei, H., Yan, B., Gagneur, J., Conradt, B. (2017) Caenorhabditis elegans CES-1 snail represses Pig-1 MELK expression to control asymmetric cell division. Genetics; 206(4), PubMedID: 28652378
Vladimir Potapov, Jennifer L. Ong. (2017) Examining Sources of Error in PCR by Single-Molecule Sequencing. PLOS One; PubMedID: 28683110
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