Dimethyl Sulfoxide (DMSO) is often included in Polymerase Chain Reaction (PCR) mixtures primarily to improve the amplification of difficult DNA templates.
Key Roles of DMSO in PCR
Based on scientific literature and practical experience, including the provided reference, DMSO serves several critical functions that can enhance PCR performance:
- Enhance Amplification: Addition of DMSO is known to directly boost the efficiency of the amplification process, leading to a stronger yield of the target DNA sequence.
- Reduce Secondary Structures: One of the main reasons for using DMSO is its ability to interfere with the formation of complex secondary structures within DNA. This is particularly important when working with templates that have a high GC content, where strong bonds can form loops and hairpin structures that hinder the polymerase enzyme. By disrupting these structures, DMSO ensures smoother primer binding and DNA synthesis.
- Relax Supercoiled Plasmids: For templates derived from supercoiled plasmids, DMSO can help relax the DNA molecule. It achieves this by increasing the prevalence of single-stranded regions. This relaxation can facilitate access for enzymes like Topoisomerase I, which might be present or become relevant in certain reaction setups, although its primary benefit in standard PCR relates to the overall DNA structure accessibility.
In summary, DMSO acts as an important additive, particularly for challenging templates, by improving DNA accessibility and ensuring efficient amplification.
When to Consider Using DMSO
You might consider adding DMSO to your PCR reaction buffer under the following circumstances:
- Amplifying templates with known high GC content (typically >60%).
- Difficulty in amplifying a target sequence despite optimization.
- Working with long DNA fragments.
- Using templates derived from supercoiled plasmids.
Typical concentrations of DMSO in PCR range from 2% to 10% (v/v), but optimization may be required for specific reactions.
