Enhanced Diagnostic Tools
Need a rapid, high accuracy and sensitive PCR methods for the detection of rare mutant DNA within an excess wild-type sample? Bio-Synthesis's BNA PCR Clamping gives you exceptional high specificity and sensitivity and also allows detection of mutants in low abundance, otherwise undetectable by other mutation-specific fluorescent probes.
BNAClampTM Technology takes full advantages of BNA probes that have strong binding affinity, specificity to its complementary strands. BNA enhanced clamp oligonucleotide can be easily apply to detect only the mutated DNA sequences from a mixture with the wild type DNA seqeunce by PCR amplification. BNA probe tightly bound to wild type sequences and blocks its PCR amplification while allowing amplification of mutant sequence of imperfect match. Single base mismatch is enough to discriminate amplification of mutant type from wild type. The short BNA DNA PCR clamp construct (10-18 mers) exhibit remarkable sensitivity of detection as low as 0.01% mutated DNA in the sample analyzed.
Contact Bio-Synthesis for sequence-specific control of gene expression by BNA PCR Clamp oligonucleotide.
The ability to identify rare mutant DNA within an excess wild type sample is laborious. Since 90% of sequence variants in human are often differ in single bases of DNA, called single nucleotide polymorphisms (SNPs). Polymorphisms within an individual may cause cancer or other diseases. Successful detection system is essential for characterizing early detection for cancer patients, genetic disease and immune response and others.
Over the past two decades, much research has focused on improving the selectivity of PCR-based technologies for enhancing the detection of minority (mutant) alleles in clinical samples. Routine application in clinical and diagnostic settings requires that these techniques be accurate and cost-effective and require little effort to optimize, perform, and analyze. However, the limitation using oligonucleotide to target duplex DNA often cause by its poor binding ability and low biostability against nuclease degradation.
Bio-Synthesis's Bridged Nucleic Acid (BNA) mediated PCR clamp and mutant-specific probes are versatile and sensitive antigene technology. It can be used to identify single nucleotide changes in DNA molecules ( < 0.01% of the total DNA). BNA clamping specifically blocks amplication of a given DNA template, while allowing amplification of another template that differs by as little as one nucleotide. BNA enhanced oligonuncleotide have properties unique form DNA that allow BNA clamping. First, BNA/DNA interactions are generally 2-4 oC per base pair more stable than the corresponding DNA/DNA duplex. The shorter probes can be designed to address traditionally problematic target sequences, such as AT- or GC-rich regions, higly repetitive sequences or regions with difficult secondary structure. Shorter regions of homology in aligned sequences can also be targeted. BNA is compatible with real time PCR system. As a result, BNA enhanced DNA/BNA PCR clamps can be used to compete with probes and primers for binding. They are very similar to the primers or probes they compete with, but are designed to perfectly match undesired PCR products or templates that should not be amplified. BNA mediated mutation-specific detection probe are simple, sensitive, an ideal solution to detect minority mutant population in the diagnosis of infectious diseases.
Suggested design for the ratio of BNA modification in the clamp oligos should be varies by type of BNA-NC analogs use, oligo length, GC%, mutation kind, Tm value etc. Generally more than 50-80 of 10-15 mer DNA oligo should be use. BNA's should be introduced at the positions where specificity and discrimination is needed (e.g. 3' end in allele specific PCR and in the SNP position in allele specific hybridization probes). 2’OMe modified base are used at 5’end for exonuclease protection The BNA/DNA clamps are designed not be be extended during the PCR reaction, we suggest oligo 3' end to be modified with a phosphate group or dideoxy terminal modifier to stop primer extension.