Helper-Dependent Chain Reaction (HDCR), a Unique PCR Method with Sensitivity and Specificity Suitable for DNA Diagnostic Assays

摘要

We have developed a unique PCR-based technique that amplifies free DNA ends by a modified priming scheme which enforces selection for the target sequence throughout every PCR amplification cycle and thus results in enhanced specificity and sensitivity. This technique, Helper-dependent Chain Reaction (HDCR), is particularly useful for the detection of minute amounts of rare target DNA in the presence of a vast excess of non-target DNA; for example, the detection of biomarkers in circulating plasma DNA. Targeted fragments are tagged using specially designed 'helper' oligonucleotides that are also used to maintain selection in subsequent amplification cycles. The process uses disabled primers called 'drivers' that can only prime on each cycle if the helpers recognize specific sequences within the target amplicon. In this way, selection for the sequence of interest is maintained throughout the amplification, preventing amplification of unwanted sequences.