DNA templates with a high proportion (>65%) of GC (guanine and cytosine) nucleotide base pairs can be difficult to amplify during PCR. This is because the strength of hydrogen bonding between these nucleobases is greater than that of the other nucleobases (adenine and thymine), and because GC-rich templates have a tendency to fold into complicated secondary structures (e.g. loops and hairpin structures). This causes GC-rich templates to be resistant to melting in the denaturation steps during PCR, preventing accurate primer binding and polymerase amplification.

10x GC-Rich PCR Enhancer is used as a PCR additive for difficult to amplify GC-rich templates. The optimised solution modifies the melting behaviour of nucleic acids, and often enhances amplification of suboptimal PCR templates with high degrees of secondary structures and GC-rich regions.

• Improve amplification success: Efficient amplification of DNA sequences with higher than 65% GC content
• Reduce PCR inhibition: High inhibitor tolerance
• Increase target detection: Reduce non-specific amplification

Additive for PCR reactions with more demanding DNA templates

Recommended Usage

10x GC-Rich PCR Enhancer should be used at a defined working concentration (1x, 2x or 3x solution), and only if non-specific amplification occurs. For a standard 20 µL reaction this would require replacing 2 – 6 µL of 10x GC-Rich PCR Enhancer respectively for PCR grade water within the reaction.

Components

10x GC-Rich PCR Enhancer.

Storage & Stability

Routine storage at –20ºC. Temporary storage at room temperature while in use has no detrimental effects on the quality of this reagent. Optionally aliquot into sterile tubes and freeze to minimise potential microbial contamination after opening.

Shipping conditions

Shipped at room temperature.