High Pure PCR Template Preparation Kit

Low to medium throughput genomic DNA isolation.

High Pure PCR Template Preparation Kit; Instructions For Use

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.

Note: A special Inhibitor Removal Buffer is included which allows the use of heparinized sample material (100 U/mL of heparin). This buffer increases the sensitivity and reproducibility of assays performed with the isolated nucleic acid, even when the sample contains heparin.

Capacity: The High Pure Spin Filter Tubes hold up to 700 μL sample volume.

High Pure PCR Template Preparation Kit has been used in the extraction of genomic DNA from whole blood samples, cervical lesion specimens, gastric muscosal tissue and cervical carcinoma cell lines.

The High Pure PCR Template Preparation Kit purifies nucleic acids from a wide variety of sample materials including whole blood, cultured cells, and tissue. The isolated nucleic acids can be used for:

• Long-template PCR
• Real-time, quantitative PCR
• SNP detection
• Southern blotting
• Cloning

The High Pure PCR Template Preparation Kit purifies nucleic acids from a wide variety of sample materials, including whole blood, cultured cells, and tissue samples. Bacteria and yeast require a specific pre-lysis treatment with lysozyme or lyticase.

• Minimize DNA loss using a kit that removes contaminants without precipitation or other handling steps that degrade DNA.
• Recover high molecular weight DNA (30 to 50 kb) that is suitable for long-template PCR.
• Improve reliability and reproducibility in downstream applications (real-time, quantitative PCR).
• Save time and maximize flexibility by preparing multiple PCR templates simultaneously.
• Eliminate the use of hazardous organic compounds such as cesium chloride, phenol, chloroform, and ethidium bromide.

DNA is isolated from 25 mg of calf thymus, 1 x 10⁶ K562 cells, and 200 μl of EDTA whole blood. Yield is measured via OD for tissue and cell samples. The quality of the nucleic acid is controlled in an Expand Long Range PCR with a 9.3 kb amplification product for DNA derived from cells. PCR on a LightCycler^® Instrument is performed on human blood research samples using kits for Factor V Leiden and CycA.

Blood cells or tissue are lysed by a short incubation with a special Lysis Buffer and Proteinase K in the presence of a chaotropic salt such as guanidine-HCl, which immediately inactivates all nucleases. Cellular nucleic acids bind selectively to special glass fibers pre-packed in the High Pure Purification Filter Tube. Bound nucleic acid is purified in a series of rapid wash-and-spin steps to remove contaminating cellular components. Finally, low salt elution releases the nucleic acid from the glass fiber. This simple method eliminates the need for organic solvent extractions and DNA precipitation, allowing for rapid purification of many samples simultaneously.

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