Simplify the Preparation of Transfection-Grade Plasmid DNA

ZymoPURE Endotoxin-Free Plasmid Purification Kits

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Ultra-Fast Midi & Maxipreps

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PROTOCOL
DATA SHEET
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Fastest

Simple 16-minute Midi/Maxi preps

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Highest Yield

Up to 25 mg from a spin-column

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Ultra-Pure

Endotoxin-Free & Transfection Ready

Endotoxin-Free Plasmid Purification Reinvented

Save more than 2 hours per prep!
Plasmid Purification
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Image of a horizontal bar graph showing the protocol length, in minutes, with ZymoPURE II only taking 16 minutes.
Simply bind, wash, and elute.

Simply bind, wash and elute. Novel binding chemistry and spin-column enables simple purification of transfection-ready plasmid DNA up to 9x faster using a vacuum manifold or centrifuge. No slow gravity flow columns, no tedious alcohol precipitation.

Highest Yield. Lowest Elution Volume.

Up to 6x More Concentrated Plasmid
Up to 3x More Plasmid DNA Yield

Yield and concentration for plasmid DNA isolated using the ZymoPURE II Maxiprep kit compared to two endotoxin-free plasmid purification kits from Supplier Q and Supplier MN. Plasmid DNA (pGL3®) was isolated from 150 ml of JM109 E. coli culture grown overnight following the manufacturer’s suggested protocol (in duplicate). One (1) µl of eluted plasmid DNA was visualized post agarose gel electrophoresis. M, ZR 1 kb DNA Marker (Zymo Research).

Transfection Ready Plasmid DNA

Transfection-grade Plasmid DNA
Transfection-grade Plasmid DNA 400x less endotoxin

Endotoxin-Free

Purify Endotoxin-Free Plasmid with ZymoPURE II Midi, Maxi & Gigaprep Kits

Transfection-grade Plasmid DNA application ready

Application-ready

CRISPR, gene modification, lentiviral vectors, synthetic biology, etc.

Transfection-grade Plasmid DNA ultra-pure

Ultra-Pure

For sensitive primary cells, in vivo injections, cloning, etc.

Superior Transfection Efficiency

HeLa Cells seeded in a 6-well plate were transfected with either 2 or 4 µg of pCI-neo®+GFP plasmid isolated from 100 ml of bacteria culture using the ZymoPURE Midiprep Kit, ZymoPURE Maxiprep Kit, or MN Midiprep Kit. GFP expressin was assessed 48 hours later in cell lysates using western blot and Ponceau S staining. The blot was also probed with an antibody against alpha-Tubulin in order to verify equal loading samples.

Data generated by V.B. at University of Cologne

Unrivaled Technology

ZymoPURE II Maxiprep Kit Supplier Q Endofree Maxiprep Supplier MN Endofree Maxiprep
Maximum Yield 3,000 µg 500 µg* 2,000 µg*
Format Spin-Column Anion-Exchange Anion-Exchange
Prep Time 16 Minutes 150 Minutes 90 Minutes
6 Minute Bind, Wash, Elute
Spin Column Elution
Alcohol Precipitation-Free
Endotoxin-Free**

*Manufacturer's stated maximum yield

**Defined as < 0.1 EU/µg of plasmid DNA

ZymoPURE endotoxin-free plasmid purification kits are the fastest and simplest plasmid purification methods available to efficiently isolate a high yield of transfection-grade plasmid DNA from E. coli in as little as 16 minutes. Plasmid DNA is endotoxin-free and ready for immediate use in downstream applications such as transfection, in vivo injections, in vitro transcription, molecular cloning, PCR, and many more.

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Cited for Sensitive Applications

Cell Therapy   |   Gene Therapy   |   Protein Replacement Therapy   |  
Recombinant Antibody Therapy   |  
Vaccine Development   |  
CAR T-Cell Therapy/Immunotherapy   |   Transfection   |  
Gene Editing   |  
Viral Vector Production (Lentivirus/Adenovirus/AAV)  |  
In vitro transcription/translation   |  
Transgenic organism generation   |   Microinjection of embryos   |  
and more…

ZymoPURE Plasmid Purification Kits

ZymoPURE Plasmid Miniprep Kit ZymoPURE 96 Plasmid Miniprep Kit ZymoPURE II Plasmid Midiprep Kit ZymoPURE II Plasmid Maxiprep Kit ZymoPURE II Plasmid Gigaprep Kit
Processing Volume ≤ 5 ml ≤ 5 ml ≤ 50 ml ≤ 150 ml ≤ 2.5 L
Processing Time ≤ 15 minutes ≤ 60 minutes ≤ 18 minutes ≤ 18 minutes ≤ 45 minutes
Yield ≤ 100 µg ≤ 100 µg ≤ 1.2 mg ≤ 3.0 mg ≤ 25 mg
Elution Volume ≥ 25 µl ≥ 125 µl ≥ 150 µl ≥ 300 µl ≥ 3 ml
Endotoxins ≤ 1 EU/µg DNA ≤ 1 EU/µg DNA ≤ 0.025 EU/µg DNA ≤ 0.025 EU/µg DNA ≤ 0.025 EU/µg DNA
Instruction Manual
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The ZymoPURE family of plasmid purification kit formats handle different amounts of culture input, ranging from miniprep (≤ 5 ml), midiprep (≤ 50 ml), maxiprep (≤ 150 ml) and gigaprep (≤ 2.5 L).

Endotoxin-Free Plasmid Purification Kits

Uses for Endotoxin-Free Plasmid Purification Kits

Use endotoxin-free plasmid purification kits to remove contaminating endotoxins during the extraction of plasmid DNA from E. coli. Endotoxins, also known as lipopolysaccharides or LPS, are a component of the cell membrane of most gram-negative bacteria. This molecule serves a practical purpose for the bacteria as it helps preserve the integrity of the cell wall and maintains a charge differential across the membrane that stabilizes the bacteria. LPSs are so integral to cell survival that the lack of it will effectively kill the cell, preventing its removal in all but a few strains of bacteria. Due to its ubiquity in gram-negative bacteria, most eukaryotic immune systems recognize endotoxins and will respond with a strong immune response. Additionally, because of the chemical structure of LPS and its tendency to form micelles, it often copurifies with DNA during traditional plasmid preparation workflows. Consequently, the presence of endotoxins in purified plasmid is a major issue for researchers working with mammalian systems, as it can significantly affect cell behavior, or even cause cell death, if introduced during transfection. Therefore, an endotoxin-free plasmid purification kit is necessary when preparing plasmid DNA for transfecting mammalian cells or tissues with plasmid DNA.

How is Endotoxin-Free Plasmid typically purified?

Traditionally, endotoxin-free plasmid was purified using Cesium Chloride (CsCl) density gradient centrifugation. However, this method requires the use of hazardous reagents and expensive ultracentrifuges. In addition, the density gradient centrifugation, phase separation, and alcohol precipitation steps are long and tedious to perform and prone to errors. Typically, a significant amount of DNA is lost during CsCl extraction. The process for purifying endotoxin-free plasmid DNA was later improved with the introduction of anion-exchange column-based endotoxin-free plasmid purification kits. These types of endotoxin-free plasmid purification kits dramatically reduced the time and number of steps it takes to prepare endotoxin-free plasmid DNA compared to CsCl density gradient centrifugation, but they typically still take several hours to perform because they rely on slow gravity flow columns to initially purify the plasmid DNA and multiple lengthy centrifugation steps to clean-up and concentrate the eluted plasmid DNA further.

We’ve pioneered rapid, efficient endotoxin-free plasmid purification kits with the introduction of our ZymoPURE Plasmid Purification product line. This family of plasmid purification kits feature a patented binding chemistry that enables simple purification of highly concentrated (up to 6 mg/ml) endotoxin-free plasmid DNA using a spin-column. The streamlined workflow eliminates slow gravity flow anion-exchange columns and isopropanol precipitation steps found in other kits. ZymoPURE plasmid purification kits reduce the time it takes to purify endotoxin-free plasmid by up to 9x using a vacuum manifold or centrifuge. Simply bind, wash, and elute transfection ready, endotoxin-free plasmid in minutes.

The ZymoPURE plasmid purification kits are optimized to ensure the eluted DNA is free of endotoxins, salt, protein, and RNA, resulting in plasmids that are suitable for use in sensitive applications. The resulting endotoxin-free plasmid DNA is ideal for transfection (including sensitive and primary cells), CRISPR-Cas9 and gene editing, lentiviral vectors, adenovirus vectors, AAV vectors, gene therapy, chimeric antigen receptor (CAR) generation, recombinant antibody generation, in vitro transcription and translation, synthetic biology, PCR, transgenic organism generation and microinjections, molecular cloning, restriction endonuclease digestions, site-directed mutagenesis, plasmid transformation of competent cells, Sanger sequencing, and other sensitive downstream applications.

Do I Need an Endotoxin-Free Plasmid Purification Kit?

Both standard and endotoxin-free plasmid purification kits will prepare plasmid DNA from E. coli culture that is suitable for a variety of downstream applications. Endotoxins will not negatively affect many routine molecular biology techniques, such as transforming bacteria and yeast, DNA sequencing, PCR, restriction enzyme digestions, ligations, gibson assembly, and golden gate assembly, so an endotoxin-free plasmid purification kit is not always necessary. However, when introducing the plasmid into a biological system that is sensitive to endotoxins, such as mammalian cells or tissues, it is imperative that an endotoxin-free plasmid purification kit is used to prepare plasmid DNA that is low in endotoxins, so the experiment and results are not comprised. Also, not all standard plasmid purification kits are created equally, and some might be more prone to the carryover of other contaminants, such as salts, proteins, and RNA, which will also be detrimental to a variety of molecular biology techniques. Therefore, since endotoxin-free plasmid purification kits have typically been optimized to produce the highest-quality plasmid, they are generally the safest bet for preparing plasmid DNA for any sensitive downstream application.

Which Endotoxin-Free Plasmid Purification Kit is right for me?

There are a couple important points to consider when selecting an endotoxin-free plasmid purification kit. First, is the amount of overnight E. coli culture you want to process and the amount of plasmid DNA you are trying to prepare, which will generally be determined by your downstream transfection experiments (number of cells to transfect, transfection method, number of replicates, etc.). Our endotoxin-free plasmid purification kits are available in a variety of scales, including mini, midi, maxi, and giga preps. Therefore, you can process just a few milliliters of overnight culture to purify tens of micrograms of plasmid with a miniprep or liters of overnight culture to purify milligrams of plasmid DNA with a gigaprep. Second, is the level of endotoxins in the prepared plasmid DNA, which will largely be determined by the biological system that you are going to transfect. An endotoxin level in the purified plasmid of less than 50 endotoxin units per microgram of plasmid (< 50 EU/µg) will generally be sufficient for transfecting standard immortalized cell lines, such as HeLa, HEK293, and CHO. However, for primary cells, it is recommended to use plasmid that has less than 10 endotoxin units per microgram of plasmid (< 10 EU/µg) and 1 endotoxin unit per microgram of plasmid (≤ 1 EU/µg) for sensitive primary cells. For in vivo transfection and transfecting immune cells, it is recommended to use purified plasmid DNA that has less than 0.1 endotoxin units per microgram of plasmid (≤ 0.1 EU/µg). Zymo Research provides endotoxin-free plasmid purification kits that are sufficient for preparing plasmid DNA for even these most sensitive transfection experiments.

ZymoPURE Citations for Sensitive Applications

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