QIAfilter Plasmid Kits

For fast purification of up to 10 mg transfection-grade plasmid or cosmid DNA

Features

  • Lysate clearing without centrifugation
  • Reduced plasmid purification time
  • High yields of plasmid DNA
  • LyseBlue reagent for optimum lysis and maximum DNA yield
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QIAfilter Plasmid Midi Kit (25)

Cat. No. / ID: 12243

25 QIAGEN-tip 100, Reagents, Buffers, 25 QIAfilter Midi Cartridges
₩498,000.00
KitCartridge
QIAfilter Plasmid Kit
QIAfilter Cartridge
Cartridge type
Midi
Maxi
Preparations
25
100
Add to cart
QIAfilter Plasmid Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

Product Details

QIAfilter Plasmid Kits provide anion-exchange-based plasmid purification with bacterial lysate clearing by filtration. Purified DNA is equivalent to that obtained by
2 x CsCl gradient centrifugation and is suitable for transfection-grade applications.

 

The QIAfilter Plasmid Mega Kit (cat. no. 12281) and the QIAfilter Plasmid Giga Kit (cat. no. 12291) will be discontinued on June 30, 2021. They remain available as long as stocks last.
The QIAGEN Plasmid Mega Kit (cat. no. 12181) and the QIAGEN Plasmid Giga Kit (cat. no. 12191) can be used with the QIAfilter Mega-Giga Cartridges (cat. no. 19781) as an optional protocol step for rapid clearing of bacterial lysates by filtration instead of centrifugation. Visit the QIAGEN plasmid kits web page to learn more.

Performance

The QIAfilter Plasmid Kits combines QIAfilter Cartridges for rapid clearing of bacterial lysates by filtration instead of centrifugation, with proven QIAGEN-tips containing anion-exchange resin for efficient plasmid purification. Yields of up to 10 mg (giga), 2.5 mg (mega), 500 µg (maxi), and 100 µg (midi) transfection-grade, high-copy plasmid DNA are achieved from culture (culture volumes depend on plasmid copy number, size of insert, host strain, and culture medium). 

 

Principle

QIAfilter Cartridges, provided in QIAfilter, HiSpeed, and EndoFree Plasmid Kits, are special filter units designed to replace centrifugation following alkaline lysis of bacterial cells. QIAfilter Cartridges completely remove SDS precipitates and clear bacterial lysates in a fraction of the time needed for centrifugation, reducing plasmid-purification time by up to 1 hour.

As an optional protocol step for rapid clearing of bacterial lysates by filtration instead of centrifugation, you can use the QIAfilter Mega-Giga Cartridges (cat. no. 19781) with the QIAGEN Plasmid Mega Kit (cat. no. 12181) and the QIAGEN Plasmid Giga Kit (cat. no. 12191). Visit the QIAGEN plasmid kits web page to learn more.
QIAfilter Maxi and Midi Cartridges use a syringe format and lysates are cleared rapidly by pushing the liquid through the filter.

The unique anion-exchange resin in QIAGEN-tips is developed exclusively for the purification of nucleic acids. Its exceptional separation properties result in DNA purity equivalent or superior to that obtained by two successive rounds of CsCl gradient centrifugation. Prepacked QIAGEN-tips operate by gravity flow and never run dry, minimizing the hands-on time required for plasmid preparation. The entire QIAGEN plasmid purification system avoids the use of toxic substances such as phenol, chloroform, ethidium bromide, and CsCl, minimizing hazard both to the user and the environment.

Specifications

Features
QIAfilter Plasmid
Maxi Kit
QIAfilter Plasmid
Midi Kit
Applications Transfection, cloning, sequencing, etc. Transfection, cloning, sequencing, etc.
Culture volume/starting material 100–250 ml culture volume 25–50 ml culture volume
Plasmid type High-copy, low-copy, cosmid DNA High-copy, low-copy, cosmid DNA
Processing Manual (filtration) Manual (filtration)
Sample per run 1 sample per run 1 sample per run
Technology Anion-exchange technology Anion-exchange technology
Time per run 120 min 110 min
Yield <500 µg <100 µg
 

 

Procedure

Neutralized bacterial lysates are incubated directly in the QIAfilter Cartridge and cleared rapidly by filtration. The cleared lysate is then loaded onto the anion-exchange tip where plasmid DNA selectively binds under appropriate low-salt and pH conditions. RNA, proteins, metabolites, and other low-molecular-weight impurities are removed by a medium-salt wash, and ultrapure plasmid DNA is eluted in high-salt buffer (see flowchart " QIAGEN Plasmid Kit procedures"). The DNA is concentrated and desalted by isopropanol precipitation and collected by centrifugation.

See figures

Applications

DNA obtained with QIAfilter Plasmid Kits provides excellent results in all applications, from cloning to transfection.

Supporting data and figures

Resources

Supplementary Protocols (2)
This protocol is for purification of up to 100 µg endotoxin-free plasmid DNA using QIAGEN-tip 100.

Publications

Comparing regional transcript profiles from maize primary roots under well-watered and low water potential conditions.
Poroyko V; Spollen WG; Hejlek LG; Hernandez AG; LeNoble ME; Davis G; Nguyen HT; Springer GK; Sharp RE; Bohnert HJ;
J Exp Bot; 2006; 58 (2):279-89 2006 Sep 21 PMID:16990373
Involvement of the pituitary-specific transcription factor pit-1 in somatolactotrope cell growth and death: an approach using dominant-negative pit-1 mutants.
Pellegrini I; Roche C; Quentien MH; Ferrand M; Gunz G; Thirion S; Bagnis C; Enjalbert A; Franc JL;
Mol Endocrinol; 2006; 20 (12):3212-27 2006 Aug 10 PMID:16901973
Participation of the Lys313-Ile333 sequence of the purinergic P2X4 receptor in agonist binding and transduction of signals to the channel gate.
Yan Z; Liang Z; Obsil T; Stojilkovic SS;
J Biol Chem; 2006; 281 (43):32649-59 2006 Sep 5 PMID:16954225
Interaction of the Akt substrate, AS160, with the glucose transporter 4 vesicle marker protein, insulin-regulated aminopeptidase.
Peck GR; Ye S; Pham V; Fernando RN; Macaulay SL; Chai SY; Albiston AL;
Mol Endocrinol; 2006; 20 (10):2576-83 2006 Jun 8 PMID:16762977
Polyamine-regulated unproductive splicing and translation of spermidine/spermine N1-acetyltransferase.
Hyvönen MT; Uimari A; Keinänen TA; Heikkinen S; Pellinen R; Wahlfors T; Korhonen A; Närvänen A; Wahlfors J; Alhonen L; Jänne J;
RNA; 2006; 12 (8):1569-82 2006 Jun 29 PMID:16809818

FAQ

What is the composition of buffer STE?

The composition of Buffer STE is:

  • 100 mM NaCl
  • 10 mM Tris-Cl, pH 8.0
  • 1 mM EDTA

Buffer STE is a DNA resuspension and storage buffer used in QIAGEN Plasmid Kits for plasmid purification and in some plasmid supplementary protocols. Details on buffer preparation and storage are presented in Appendix B of the QIAGEN Plasmid Purification Handbook.

FAQ ID -415
What is the advantage of running an analytical gel with fractions of my plasmid preparation?

Running fractions saved from each step in the plasmid preparation procedure on an agarose gel enables monitoring the performance of each crucial step in the protocol. If the plasmid DNA is of low yield or quality, the samples can be analyzed to determine at what stage of the purification procedure the difficulty occurred.

Aliquots can be taken from the cleared lysate and the flow-throughs as indicated in the relevant protocols, precipitated with isopropanol and resuspended in a small volume of TE buffer.

Please see the Troubleshooting Section of the QIAprep Miniprep Handbook and Appendix A of the QIAGEN Plasmid Purification Handbook for instructions, and a picture and legend explaining the typical results you may see. You can also access this information on our Plasmid Resource Pages.

FAQ ID -769
What is the composition of buffer QF?

Buffer QF is the elution buffer used in QIAGEN Plasmid Kits for plasmid purification and in QIAGEN Blood & Cell culture kits.

The composition of Buffer QF is:

  • 1.25 M NaCl
  • 50 mM Tris-Cl, pH 8.5
  • 15% isopropanol (v/v)

 

To make 1 liter of solution, dissolve 73.05 g NaCl and 6.06 g Tris base in 800 ml distilled water. Adjust the pH to 8.5 with HCl. Add 150 ml pure isopropanol. Adjust the volume to 1 liter with distilled water. Store at 15–25°C

FAQ ID -413
Which QIAGEN plasmid preparation kits will contain LyseBlue Reagent?

LyseBlue reagent is provided in the following QIAGEN plasmid kits:

FAQ ID -865
What is the recipe for LB?
Preparation of LB medium: Dissolve 10 g tryptone, 5 g yeast extract, and 10 g NaCl in 800 ml dH2O. Adjust the pH to 7.0 with 1 N NaOH. Adjust the volume to 1 liter with dH2O. Sterilize by autoclaving.
FAQ ID -212
How do I know if my plasmid is a high- or low copy number type?

Find out which origin of replication your plasmid contains, and look at the table below for classification into high-copy or low-copy types. This table can also be found online at the QIAGEN Plasmid Resource Center in the section 'Growth of bacterial cultures; Plasmid Copy Number' . A way to determine experimentally if the copy number of your plasmid is high or low is to perform a miniprep. A high-copy plasmid should yield between 3-5 ug DNA per 1 ml LB culture, while a low-copy plasmid will yield between 0.2-1 ug DNA per ml of LB culture.

 

Origins of replication and copy numbers of various plasmids and cosmids

DNA construct Origin of Replication Copy number Classification
Plasmids      
pUC vectors pMB1* 500–700 high copy
pBluescript® vectors ColE1 300–500 high copy
pGEM® vectors pMB1* 300–400 high copy
pTZ vectors pMB1* >1000 high copy
pBR322 and derivatives pMB1* 15–20 low copy
pACYC and derivatives p15A 10–12 low copy
pSC101 and derivatives pSC101 ~5 very low copy
Cosmids      
SuperCos pMB1* 10-20 low copy
pWE15 ColE1 10-20 low copy

* The pMB1 origin of replication is closely related to that of ColE1 and falls in the same incompatibility group. The high-copy plasmids listed here contain mutated versions of this origin.

FAQ ID -350
Can I eliminate RNase A from buffer P1 for my plasmid preparation to obtain RNase-free DNA for in-vitro transcription?
No, RNase A should not be omitted from buffer P1. It is required to prevent RNA contamination of the purified plasmid DNA. RNase A will not interfere with downstream in-vitro transcription experiments, since it will be efficiently removed during the plasmid purification procedures using QIAGEN Plasmid Kits.
FAQ ID -366
I left Buffer P1 at room temperature after addition of RNase A, what shall I do?

Buffer P1 with RNase A used in QIAGEN Plasmid Purification Kits should be fine at room temperature for a few days. We would expect the enzyme to have some residual activity. However, optimal results cannot be guaranteed after storage at room temperature. If you notice that RNase A activity is substantially reduced, you can add fresh RNase A to your buffer.

We recommend that Buffer P1 with RNase A be stored in the refrigerator (2-8°C). RNase A will be stable for 6 months under this condition.

FAQ ID -859
What is the composition of Buffer P2?

The composition of Buffer P2 is:

  • 200 mM NaOH
  • 1% SDS (w/v)

It should be stored at room temperature. Buffer P2 is the lysis buffer used in a variety of QIAGEN kits for plasmid DNA purification. Details on buffer preparation and storage are presented in Appendix B of the QIAGEN Plasmid Purification Handbook.

FAQ ID -203
With LyseBlue reagent for lysis control, can I now process more bacterial culture and overload the columns?

No. The maximum culture volumes recommended for QIAGEN's plasmid preparation kits still apply, and should be strictly followed. LyseBlue reagent now allows the user to monitor potential problems (insufficient bacterial cell resuspension and lysis as a consequence of overloading) early in the plasmid preparation process.

FAQ ID -864
What are the additional plasmid bands I see on my gel?

Open circular plasmid, resulting from single strand nicks, usually migrates slower in agarose gels and forms (faint) bands above the supercoiled plasmid DNA band. Sometimes an additional band of denatured supercoiled DNA migrates just below the supercoiled form. This form may result from prolonged alkaline lysis with Buffer P2 and is resistant to restriction digestion.

For a detailed description on how to run and interpret an analytical gel, please see Appendix A in the QIAGEN Plasmid Purification Handbook: "Agarose Gel Analysis of the Purification Procedure", or visit this link.

FAQ ID -1059
How can I increase low-copy plasmid DNA yields using QIAfilter Plasmid Kits?

If low-copy plasmids are used with QIAfilter Plasmid Kits, the cell lysates of two QIAfilter Cartridges can be filtered into one previously equilibrated QIAGEN-tip, resulting in doubled DNA yields.

Similarly, if low copy plasmids are used with HiSpeed Plasmid Kits, the lysates of two QIAfilter Cartridges can be filtered into one equilibrated HiSpeed Tip.

FAQ ID -1062
What can I do when the DNA pellet prepared with QIAGEN Plasmid Kits has been overdried?
Redissolve the DNA by warming the solution slightly, e.g. incubate it at 37°C, and allow more time for redissolving.
FAQ ID -572
Can I use a QIAfilter Cartridge for purifying large plasmids?
Data from QIAGEN laboratories show that cosmids of up to 45 kb can be purified using the QIAfilter procedure without any detectable DNA shearing (see figure below).


Note: Many large plasmids are present in very low copy number in cells. For very low-copy-number constructs, refer to the special protocol 'Very Low-Copy Plasmid/Cosmid Purification using QIAGEN-tip 100 or QIAGEN-tip 500 in the QIAGEN Plasmid Purification Handbook. For purification of constructs larger than 50 kb (up to 250 kb) we recommend using the QIAGEN Large-Construct Kit.
FAQ ID -121
How can I increase DNA concentration using QIAprecipitators of the HiSpeed Plasmid Kits?
Use 500 µl instead of 1 ml of Buffer TE for elution of plasmid DNA from the QIAprecipitator of the HiSpeed Plasmid Kits. If low copy plasmids are used, the lysates of two QIAfilter Cartridges can be filtered into one equilibrated HiSpeed Tip. HiSpeed Maxi Kits will result in higher DNA concentration than HiSpeed Midi Kits, because the QIAprecipitators in both kits (Midi- and Maxi Module) use the same elution volume.
FAQ ID -1061
Where can I find a protocol for cleanup of already purified plasmid DNA?

When using the silica-based QIAprep Spin Miniprep Kit, a protocol is contained in the QIAprep Miniprep Handbook, in Appendix C: Special Applications. The protocol is called: 'Purification of plasmid DNA prepared by other methods'.

For our anion-exchange based Plasmid Purification Kits, a protocol can be accessed online at our Plasmid Resource Center, and is called 'Re-Purification of Plasmid DNA Prepared by Methods other than QIAGEN Tips'.

 

FAQ ID -1031
What is the composition of buffer TE?

The composition of Buffer TE is:

  • 10 mM Tris·Cl, pH 8.0
  • 1 mM EDTA

Buffer TE is a commonly used DNA resuspension and storage buffer. It is supplied in QIAGEN's Endofree Plasmid Kits, and used for plasmid DNA resuspension in combination with other QIAGEN Plasmid Kits. Details on buffer preparation and storage are presented in Appendix B of the QIAGEN Plasmid Purification Handbook.

FAQ ID -416
What is the composition of buffer FWB2?

The composition of Buffer FWB2 is:

  • 1 M potassium acetate, pH 5.0

Buffer FWB2 is the QIAfilter wash buffer used in QIAfilter Plasmid Kits for plasmid purification. Details on buffer preparation and storage are presented in Appendix B of the QIAfilter Plasmid Purification Handbook.

FAQ ID -417
What to do if cell clumps are present after Buffer P2 addition when using LyseBlue Reagent?

If cell clumps are present after adding Buffer P2 to your samples when using a QIAGEN Plasmid Purification Kit in combination with LyseBlue Reagent, you have two options:

  • pipet the cell clumps up and down for resuspension
  • transfer any clumps to a separate tube, add Buffer P1 and mix vigorously for resuspension, add Buffer P2 for lysis, and subsequently transfer the lysed material back to combine it with the rest of the original solution

Note: Avoid incubation times longer than 5 minutes in Buffer P2 as this will irreversibly denature plasmid DNA. In the scenario above, Buffer P3 may need to be added to portions of the sample, which can be subsequently combined once resuspension, lysis and neutralization of all fractions is complete.

FAQ ID -861
What is the RNase A concentration and composition of Buffer P1?

The composition of Buffer P1 is:

  • 50 mM Tris·Cl, pH 8.0
  • 10 mM EDTA
  • 100 µg/ml RNase A

After RNase A addition, the buffer should be stored at 2–8°C.

Buffer P1 is the resuspension buffer used in a variety of QIAGEN kits for plasmid DNA purification. Details on buffer preparation and storage are presented in Appendix B of the QIAGEN Plasmid Purification Handbook.

FAQ ID -198
Can Buffers N3 and P3 be used interchangeably?
No. Although both Buffers N3 of the QIAprep Spin Miniprep and P3 of the QIAGEN Plasmid Kits perform the neutralization step in an alkaline lysis procedure, they are completely different. Buffer N3 contains a proprietary formula that sets up binding conditions for the QIAprep Miniprep column's silica-gel-membrane. Buffer P3 sets up binding conditions for QIAGEN anion-exchange columns. Our website explains QIAGEN's nucleic acid purification technologies in more detail.
FAQ ID -310
What is the composition of buffer P3?

The composition of Buffer P3 is:

  • 3.0 M potassium acetate, pH 5.5

Buffer P3 is the neutralization buffer used in QIAGEN's anion-exchange based Kits for plasmid preparation. Details of buffer preparation and storage are presented in Appendix B of the QIAGEN Plasmid Purification Handbook.

FAQ ID -418
What is the recipe for SOC medium?

The components of the SOC medium are:

  • 0.5% Yeast Extract
  • 2% Tryptone
  • 10 mM NaCl
  • 2.5 mM KCl
  • 10 mM MgCl2
  • 10 mM MgSO4
  • 20 mM Glucose*

*Note: add Glucose after autoclaving the solution with the remaining ingredients, and letting it cool down. Sterilize the final solution by passing it through a 0.2 µm filter.

SOC medium can be stored at room temperature and is stable for several years.

FAQ ID -798
What is the white insoluble precipitate in my resuspended plasmid DNA pellet?

White insoluble material in the resuspended plasmid DNA pellet indicates carry-over of salts and/or carbohydrates. Ensure that isopropanol is used at room temperature for precipitation. Some bacterial strains, such as TG1 and JM100, naturally produce a high level of carbohydrates. However, carbohydrate contamination may also be observed when using other strains. The most common cause of this problem is over-growth of bacterial cultures. To avoid this, closely follow the guidelines for Plasmid DNA Preparation in the Handbook that was provided with the respective QIAGEN Plasmid Kit.

Useful hints and information on optimizing plasmid preparations can be found at the QIAGEN Plasmid Resource Center.

FAQ ID -352
How should QIAGEN Plasmid Purification Kits be stored and for how long?
QIAGEN-tips and QIAfilter Cartridges should be stored dry and at room temperature (15–25°). They can be stored for at least two years without showing any reduction in performance, capacity, or quality of separation.

QIAGEN, QIAfilter, EndoFree, HiSpeed and Large-Construct Plasmid Kits should be stored at room temperature (15–25°). After adding RNase A, Buffer P1 should be stored at 2–8°C and is stable for six months. All other buffers and RNase A stock solution can be stored for two years at room temperature (15–25°).
FAQ ID -192
Is plasmid DNA purified with QIAGEN Plasmid Purification Kits suitable for in vitro transcription?

Plasmid preparations are free of any detectable proteins or other contaminants when purified using QIAGEN's anion-exchange kits according to the recommended protocols. DNA purified using QIAGEN Plasmid Kits, QIAfilter Plasmid Kits, or EndoFree Plasmid Kits gives excellent results with in-vitro transcription experiments.

Although a high level of RNase A is employed at the beginning of the procedure, it is removed efficiently by potassium dodecyl sulfate precipitation and subsequent washing with Buffer QC. It is possible, although not necessary, to omit RNase A from the procedure when purifying DNA for in vitro transcription. In this case, increasing the volume of Wash Buffer QC is recommended (e.g., for a Midi preparation on a QIAGEN-tip 100, use at least 2x 30 ml of Buffer QC instead of 2x 10 ml).

FAQ ID -1
What is the composition of buffer QC?

Buffer QC is the wash buffer used in QIAGEN Plasmid Kits for plasmid purification and in QIAGEN Blood & Cell Culture kits.

The composition of Buffer QC is:

  • 1.0 M NaCl
  • 50 mM MOPS, pH 7.0
  • 15% isopropanol (v/v)

 

To make 1 liter of solution, dissolve 58.44 g NaCl, 10.46 g MOPS (free acid) in 800 ml distilled water. Adjust the pH to 7.0 with NaOH. Add 150 ml pure isopropanol. Adjust the volume to 1 liter with distilled water. Store at 15–25°C.

FAQ ID -412
How can I prevent clogging of QIAfilter cartridges?
It is important to completely mix and lyse bacterial cells with plasmid Buffers P1 and P2. Incomplete mixing results in sticky or slimy areas of lysate, which will clog the filter matrix. It is recommended to completely resuspend cells in Buffer P1 and vigorously mix after addition of Buffer P2. Also mixing after Buffer P3 addition needs to be complete to allow fluffy precipitation of cell debris, which will float up. If the white debris does not float, dislodge it from the QIAfilter barrel wall (e.g. using a sterile pipette tip). Otherwise it will collect on the filter matrix and can lead to clogging. Use of LyseBlue reagent will help to achieve proper mixing results.
FAQ ID -1060
What is the composition of buffer QBT?

Buffer QBT is the equilibration buffer used in QIAGEN Plasmid Kits for plasmid purification and in QIAGEN Blood & Cell culture kits.

The composition of Buffer QBT is:

  • 750mM NaCl • 50 mM MOPS, pH 7.0
  • 15% isopropanol (v/v)
  • 0.15 % Triton® X-100 (v/v)

To make 1 liter of solution, dissolve 43.83 g NaCl, 10.46 g MOPS (free acid) in 800 ml distilled water. Adjust the pH to 7.0 with NaOH. Add 150 ml pure isopropanol and 15 ml 10% Triton X-100 solution (v/v). Adjust the volume to 1 liter with distilled water. Store at 15–25°C

FAQ ID -411
When is chloramphenicol amplification of plasmids performed?

When low-copy-number plasmids containing the pMB1 or ColE1 origin of replication are prepared using QIAGEN Plasmid Purification Kits, plasmid DNA yields can be improved by adding chloramphenicol to the culture medium (170 mg/liter) to amplify copy numbers. For details and precautions on the use of chloramphenicol when culturing plasmids, please refer to standard manuals for cloning procedures (e.g., Molecular cloning : A laboratory manual / T. Maniatis, E.F. Fritsch, J. Sambrook).

Cultures of bacteria containing low-copy-number plasmids amplified in the presence of chloramphenicol should be treated as if they contain high-copy-number plasmids when choosing the appropriate culture volumes for the QIAGEN-tip to be used.

Note that copy numbers of the current generation of plasmids are so high that selective amplification in the presence of chloramphenicol is not necessary to achieve high yields. Please see FAQ 350 for origins of replication and copy numbers of various plasmids.

FAQ ID -3
Why would clumps occur following the addition of Buffer P2 when using LyseBlue Reagent in a plasmid preparation?

Clumps that occur after addition of Buffer P2 in a bacterial lysate containing LyseBlue reagent indicate poor resuspension of the bacterial cell pellet in Buffer P1. This handling error leads to inefficient cell lysis, and incomplete precipitation of SDS, cell debris, and genomic DNA. When resuspending the cell pellet, vortexing longer or resuspending the pellet by pipetting up and down can help.

If cells have been resuspended properly in P1, “brownish areas” after P2 addition just indicate poor mixing of P1 and P2. To overcome this, continue mixing the solution by inverting it gently until a homogeneous blue suspension is achieved.

FAQ ID -862