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PCR PURIFICATION KIT - Column Type

ORDERING INFORMATION

Catalog #

Product

Amount

Price (EUR)

# PCR 50
ROVALAB PCR Purification Kit
50 preparations

43,50

# PCR 250
ROVALAB PCR Purification Kit

250 preparations

176,50

PRODUCT COMPONENTS

Buffer

# PCR 50

# PCR 250

Binding Buffer

30 ml

150 ml

Column Activation Solution

6 ml

30 ml

Elution Buffer

6 ml

30 ml

Spin Columns & 2ml Tubes

50

250

DESCRIPTION

ROVALAB PCR Purification Kit is designed for the work-up of PCR reactions (removal of primer dimers, primers, nucleotides, proteins, salt, agarose, ethidium bromide and other impurities). The preparation is based on a silica-membrane technology for binding DNA in high-salt and elution in low-salt buffer. The kit provides a simple and efficient way to purify linear or circular DNA in the size range from 100 bp to 10 kb and is optimized for working with DNA amounts of up to 20 μg. It requires no organic extractions or precipitation and guarantees high and stable recovery rates.

FEATURES

  • High Yield: > 80%
  • Easy removal of primer-dimer
  • Spin column type
  • Purification size: 100 bp - 10 kb
  • DNA amounts up to 20 g
  • Sequencing grade

APPLICATIONS

  • General sequencing
  • PCR
  • Cloning
  • Restriction enzyme digestion

BASIC PROTOCOL

The DNA purification follows a simple binding, washing and eluting procedure.

Before start, prepare an 80% Ethanol (V/V) solution as Washing buffer:

  • for 50 preparations (# PCR 50): 80 ml
  • for 250 preparations (# PCR 250): 400 ml
The additional use of Isopropanol (not included) is recommended for fragments smaller than 200 bp or larger than 5 kbp.The optional secondary washing step minimizes the salt content of the purification product but may significantly reduce the yield of DNA fragments <200 bp.

1a. Standard Sample Preparation

For DNA fragment sizes in the range of 200 bp to 5 kbp:

  • Add 5 volumes of Binding Buffer to 1 volume of DNA sample and mix well. For example, if the volume of your DNA sample is 50 μl, add 250 l Binding Buffer.

1b. High Yield Sample Preparation

For DNA fragment sizes smaller than 200 bp or larger than 5 kbp:

  • Add 3 volumes Binding Buffer and 2 volumes of Isopropanol to the PCR sample. For example, if the volume of your DNA sample is 50 μl, add 150 μl Binding Buffer and 100 l Isopropanol.

2. Column Activation

  • Place a Spin Column into a 2 ml collection tube.
  • Add 100 l of Column Activation Solution into the Spin Column.
  • Centrifuge at 10,000 g for 30 sec in a microcentrifuge.

3. Column Loading

  • Apply the sample mixture from step 1 into the activated Spin Column.
  • Centrifuge at 10,000 g for 30 sec in a microcentrifuge.
  • Discard the flow-through.

4. Column Washing

  • Place the DNA loaded Spin Column into the used 2 ml tube.
  • Apply 700 l of Washing Buffer (80% Ethanol) to the Spin Column.
  • Centrifuge at 10,000 g for 30 sec and discard the flow-through.
  • Optional Secondary Washing: Recommended only for DNA >200 bp, if highly purified DNA (for DNA sequencing, transfection etc.) is required.
    • Add 700 l of Washing Buffer to the Spin Column.
    • Centrifuge at 10,000 g for 30 sec and discard the flow-through.
  • Centrifuge again for 2 min to remove residual Washing Buffer.

5. Elution

  • Place the Spin Column into a clean 1.5 ml microtube (not provided in the kit).
  • Add 30-50 μl Elution Buffer to the center of the column membrane.
  • Incubate at room temperature for 1 min.
  • Centrifuge at 10,000 g for 1 min to elute DNA.

STORAGE

Store at room temperature (15 - 25 C) for 12 months.