New type of PCR primer that offers exquisite discrimination of single nucleotide polymorphisms

Life Sciences : Research Tools

Available for licensing

Inventors

  • Andrew Ellington, Ph.D. , Molecular Biosciences
  • Michelle Byrom , University of Texas at Austin
  • Yu Sherry Jiang , University of Texas at Austin
  • Sanchita Bhadra , Molecular Genetics and Microbiology

Background/unmet need

Accurate detection of single nucleotide polymorphisms (SNPs) is crucial for diagnosing diseases, assessing disease risk, drug efficacy and side effects, and for disease management.

Real-time polymerase chain reaction (PCR) is currently widely used for rapid and sensitive SNP diagnostics; however, there are at least two drawbacks of this method using conventional PCR primers: first, the presence of impurities or contaminants in prepared samples can lead to non-specific amplification; and second, differences in amplification efficiency from wild-type templates compared to SNP-containing templates are often too small to achieve an ideal level of discrimination.

Thus, there is a need for robust discrimination between SNPs, irrespective of sample origin, condition, preparation, or purity.

Invention Description

The Ellington lab at UT Austin has developed a toolkit of unique primers and primer design rules that distinguish single nucleotide polymorphisms with an exquisite degree of discrimination (i.e., up to 100,000-fold), resulting in little to no false positives.

The rationally designed primers contain unique structural components and a powerful mechanism of action that allow hybridization to matched targets with great efficiency while greatly diminishing binding to non-matching targets. This establishes a large amplification bias in favor of the matched template versus the non-matching template, allowing exquisite allelic distinction in real-time.

Additionally, inexpensive primers for different SNPs can be multiplexed and can lead to definitive identification of each SNP allele, even in parallel.

Benefits/Advantages

  • Can be used for the identification of SNPs in any context--identifying, for example, disease susceptibility, drug-resistant alleles of infectious organisms, or drug-resistant cancer cells to guide therapy
  • Can be used to analyze SNPs from complex mixtures of genetic material
  • Can be used in any PCR application to provide a high degree of target discrimination
  • Better discrimination than the current competitors on the market
  • Digital level of accuracy (up to 100,000-fold discrimination; i.e., yes/no level of accuracy)
  • Little to no false positives (i.e., no amplification of unmatched templates, or up to 30 cycles difference between average Cq values for wild-type and SNP containing templates)
  • Reduced primer-dimer formation
  • The presence of all possible SNPs at one site can be tested simultaneously.
  • Primers are relatively cheap and easy to manufacture and do not require specialized nucleotides.
  • May be useful for some forensic analyses; e.g., phenotypic prediction and paternity testing

Market potential/applications

Forensics, diagnostics, SNP analysis

Development Stage

Lab/bench prototype

IP Status

  • 1 U.S. patent application filed