DNA sequencing through fluorophore-free Raman spectroscopy of nucleotides
DNA sequencing through fluorophore-free Raman spectroscopy of nucleotides. This technology is a DNA sequencing system designed to differentially label nucleotides for detection by Raman spectroscopy as they are incorporated into a growing DNA molecule.
Unmet Need: A highly sensitive tagging system for nucleotide detection
Current DNA sequencing methods involve the attachment of fluorophores to each nucleotide and recording of the fluorescence signal that is produced upon excitation. This is a costly approach due to the complex amplification and analysis required. It also introduces a limit on read length, as errors in the amplification process can cause multiple copies to desynchronize after a few dozen bases. Alternative approaches that avoid the need for amplification are also limited by the required use of signal-amplifying structures, which hinder throughput. Next generation DNA sequencing methods typically increase throughput and coverage by decreasing the amount of DNA template used per reaction. The natural conclusion of this trend is to sequence single molecules of DNA or RNA individually. However, it is often difficult to detect signals produced by single nucleotide additions due to the limits of current fluorescent probes and instrumentation.
The Technology: DNA tagging method with high signal strength through Raman spectroscopy
This technology makes use of surface-enhanced Raman spectroscopy to achieve accurate, high throughput DNA sequencing. By tagging each nucleotide with special molecules that have unique Raman signatures, each nucleotide yields a distinct, unambiguous Raman scattering pattern. The use of Raman tags allows for direct detection of nucleotide incorporation at the single molecule level, without the need for amplification and with no limits imposed on throughput. Additionally, this technique is highly adaptable and can be used with most sequencing applications currently available.
The viability of the modified nucleotides as synthetic material has been demonstrated through detection of synthesized DNA products as well as through direct observation of the Raman signal associated with synthesis.
Applications:
- Cost-effective DNA sequencing
- Adaptable tagging scheme can be applied to various single molecule techniques involving nucleic acids
- Can be used to make Raman-active nucleotide triphosphate analogs, which in turn are useful in the study of motor proteins and other ATP-hydrolyzing proteins
- Other types of molecular detection applications, such as HPLC or ELISA
- Portable chemical detection devices
Advantages:
- Intensity of Raman signal can be adjusted by adding more tags to a molecule
- Does not overlap with spectra of most commonly used fluorescent probes
- Can be visualized more readily than other non-amplified single-molecule sequencing reactions
- Does not require error-prone amplification, and therefore offers greater read length
- Highly adaptable and can be integrated into existing products or developed into a novel-sequencing platform
- Each nucleotide can be labeled distinctly and unambiguously
Lead Inventor:
Patent Information:
Related Publications:
Tech Ventures Reference:
IR CU13071
Licensing Contact: Cynthia Lang