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Picture of High Pure PCR Cleanup Micro Kit

High Pure PCR Cleanup Micro Kit

For life science research only. Not for use in diagnostic procedures.

Product No. Pack Size List Price Your Price Quantity
04983955001
50 purifications $ 120.00

Product No. Pack Size List Price Your Price Quantity
04983912001
200 purifications $ 412.00

The High Pure PCR Cleanup Micro Kit efficiently purifies products from PCR and other reactions. The kit eliminates primers, mineral oil, salts, unincorporated nucleotides, and thermostable DNA polymerases, which may inhibit subsequent enzymatic reactions such as labeling, sequencing, or cloning of PCR products.

  • Conserve resources and save time
    by using a single kit with a simple, rapid protocol.
  • Obtain purified product in a small elution volume
    (≤10 μL) for demanding downstream applications.
  • Generate contaminant-free DNA
    for direct use in cloning, ligation, restriction digests, and other reactions.
  • Selectively isolate specific DNA fragment sizes
    by using the kit’s Binding Enhancer to adjust purification stringency (see Figure 1).
  • Eliminate the use of hazardous organic compounds
    such as cesium chloride, phenol, chloroform, and ethidium bromide.

 

Contents

 

  1. Binding Buffer
  2. Binding Enhancer
  3. Wash Buffer Concentrate
  4. Elution Buffer
  5. High Pure Micro Filter Tubes
  6. Collection Tubes

 

The High Pure PCR Cleanup Micro Kit is used to isolate PCR products from amplification and other reactions, and can be used in many molecular biology applications:

  • Restriction enzyme digests
  • Alkaline phosphatase treatment
  • Kinase reactions
  • Nonradioactive labeling

The kit can also be used to:

  • Purify cDNA
  • Concentrate dilute nucleic acid solutions
  • Recover DNA from agarose gel slices

Use one kit for a variety of applications.
The fast and simple High Pure protocols use a tabletop centrifuge to bind, wash, and elute the reaction product down to 10 µL (micro format) in as little as 10 minutes. The procedure conveniently eliminates a concentration step, and is ideal for downstream applications such as labeling, sequencing, cloning, ligation, or amplification using PCR. The purified DNA can also be used for Southern blotting and in vitro transcription.


 


Choose the right product for DNA fragment purification.


 

High Pure PCR Product Purification Kit

High Pure PCR Cleanup Micro Kit

Purification from agarose gel slices

X

 

Purification from reaction mixes

X

X

Concentration in 50 µL

X

 

Concentration in 10 µL

 

X

Depletion of primer-dimers <100 bp

 

X

DNA fragments up to 50 kb

X

 

DNA fragments up to 5 kb

 

X

 

A 341 bp PCR fragment of the tPA gene was amplified according to a standard block cycler protocol. The resulting reaction mixes were pooled and purified with the High Pure PCR Cleanup Micro Kit. Different amounts of Binding Enhancer were used in the purification procedure. Portions of the PCR product (250 ng each, lanes 2 – 5) and the PCR product mix (16 µL each, lanes 6 – 7) were analyzed on a 1% agarose gel (see Figure 1).
The yield from each purification is shown in Table 1.

High Pure PCR Cleanup Micro Kit, 1% agarose gel electrophoresis of 341 bp PCR product recovered in the presence of different amounts of Binding Enhancer

Figure 1:
1% agarose gel electrophoresis of 341 bp PCR product recovered in the presence of different amounts of Binding Enhancer.

Lane 1: Molecular weight marker VI
Lane 2: 0% Binding Enhancer
Lane 3: 10% Binding Enhancer
Lane 4: 20% Binding Enhancer
Lane 5: 40% Binding Enhancer
Lane 6: PCR without purification
Lane 7: PCR negative control (PCR without template)
Lane 8: Molecular weight marker VI



Binding Enhancer (%) Ratio 260/280 Yield (μg)

0

1.9

0.6

10

1.9

0.9

20

1.8

1.2

40

1.9

1.7


Table 1: Yield of DNA fragments isolated in the presence of increasing amounts of Binding Enhancer. Increasing the amount of Binding Enhancer in the isolation procedure increases the recovery of DNA fragments from the PCR product mix. As shown in lanes 2-5, however, this increase mainly reflects the differing amounts of smaller PCR fragments (e.g ., primer-dimers) co-purified with the PCR product.

Nucleic acids bind to the surface of the glass fiber fleece in the presence of a chaotropic salt (guanidine HCl). This allows the High Pure filter tube to specifically immobilize nucleic acids (both DNA and RNA) while they are freed of contaminants. The amount of DNA recovered is dependent on the amount of DNA applied to the glass fiber fleece, the elution volume, and the length of the amplification/DNA products. When 5 - 25 μg DNA is applied to the kit’s High Pure Micro Filter Tube, approximately 85% of the DNA can be recovered.

Capacity of High Pure Micro Filter Tubes: The High Pure Micro Filter Tubes hold up to 500 μL sample volume.

Typical DNA Recovery
 

Starting Material Yield/ Recovery Time Required Number of Reactions
Nucleic acids from PCR, modifying, labeling, restriction digestion reactions, and agarose gel slices 85% recovery up to 20 µg DNA 10 minutes 50 or 200


Principle

Nucleic acids bind specifically to the surface of glass fibers in the presence of chaotropic salts. Since the binding process is specific for nucleic acid, the bound material can be separated and purified from impurities by a simple wash step. The Binding Enhancer enables the modification of DNA fragment size exclusions. Small oligonucleotides and dimerized primers from amplification reactions are selectively removed. The nucleic acids elute from the glass fiber fleece in a low-salt buffer or water.

High Pure PCR Cleanup Micro Kit, principle


Quality

Greater than 70% recovery is obtained when 3 μg Roche DNA Molecular Weight Marker VI is applied to the kit’s High Pure Micro Filter Tubes. Gel electrophoresis of the eluate after purification confirms the complete removal of primer-dimers. The eluted, purified DNA shows no inhibition of amplification using a LightCycler® Instrument with the LightCycler® FastStart DNA MasterPLUS SYBR Green I.

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