Modified Nucleic Acids Research

Celadon LNA Rules (Locked Nucleic Acid design rules)

Research Papers

Position-dependent effects of locked nucleic acid (LNA) on DNA sequencing and PCR primers

Sequence-Dependent Thermodynamic Parameters for Locked Nucleic Acid (LNA)-DNA Duplex Formation

Celadon LNA Rules
Critical to DNA Nanotechnology

Background
Custom Oligonucleotide Manufacturer (COM, a fictional company) has licensed Locked Nucleic Acids (LNA) from Exiqon for custom manufacture, as well as development of catalog assays and diagnostic kits. COM anticipates the premium it can charge for LNA will lead to a substantial, highly-profitable, and enduring revenue stream.

Problem
There is no adequate software for the design of LNA primers and probes. The impediment is not just the ability to develop software, but having the knowledge of where, when, and how to place LNA for best, cost-effective results. LNA is expensive, and customers can't afford the typical primer and probe strategy of trial and error.

Solution
COM licenses Celadon LNA Rules with a co-branded version of Celadon ProbeITy.

Outcome
LNA Rules are fully integrated into the software. Now, the availability of useful software for LNA drives increased LNA use in the market. Sales and profits soar.

Moreover, LNA is found to be highly useful to the emerging market of DNA-based nanosensors. Modification of ProbeITy to design nano-structures has a meaningful impact on developing the nascent market, and LNA proves crucial to nanosensor performance. Combined, these efforts enable COM to maintain and increase its market leadership position.

Applicable Markets
As declining NextGen Sequencing costs extend the range of research and medical tests, more applications will come up against the limitations of standard methods. Probe and primer design software with LNA Rules will be key to solving problems from relatively simple structure resolution to DNA-based nanotechnology detection for biodefense.

Using its Mod-NA technology, Celadon has generated the industry's most extensive and accurate database of thermodynamic rules for precise location of LNA in primers and probes.

Applications: Research, Diagnostics, Multiplex, Pathogen Identification, Personal Genomics, Companion Diagnostics

Initial Methods: Many hundreds of published papers, Primers and Probes, microRA Analysis, RNA interference Therapeutics

Future Methods: RNA, Diagnostic Kits, DNA Nanotechnology, Food Testing, AgBio, RNA Interference Therapeutics

Celadon has done more original research on LNA:DNA thermodynamics and positioning than any other organization in the world, with nearly $1 million of state and federal research contracts and grants and more than five years invested in its scientific focus on determining hybridization and positional design rules for modified nucleic acid chemistries. Since 2001 it has conducted original research in the area of LNA thermodynamics, with the goal of better understanding LNA's melting properties and better designing DNA or RNA assays that incorporate LNA bases.

This research into applying modified nucleic acid technology to one or all of the oligonucleotides in an assay, supported by NIH and State of Maryland, has created unique, accurate databases of thermodynamic values that predict hybridization. This enables the most effective LNA design, at all possible positions: perfect match, single internal mismatch, 3' end, 5' end, multiple incorporations. Incorporable into ProbeITy, these databases rely on novel empirical methods developed by Celadon which are applicable to any modified chemistry. This represents potential value for vendors of custom LNA and SiLNA oligonucleotides, providers of multiplex platforms for for LNA incorporation, resellers of probe libraries (such as Exiqon's), and developers of pre-designed research assays and diagnostics; LNA is highly relevant because biotechnology applications demand performance that exceeds the hybridization capability of native DNA.

• Custom oligonucleotide synthesis companies can:
  • Increase sales of high-profit specialty chemicals.
• Point-of-care and other diagnostic device or assay developers can:
  • Develop research and diagnostic assays faster, with less optimization and cost.
  • Save component cost due to optimal design rules that result in least number of expensive LNAs.
  • Increased cost of LNA is trivial compared to total cost of kit.
• Developers of pre-designed research assays can:
  • Develop assays faster, with less optimization and cost.
• Large pharmaceutical, biotechnology, and other life science institutions can:
  • Improve consistency and quality.
  • Lower the cost of research and development by increasing assay design success rate, signal strength, and signal consistency.
• Academic and industry researchers can:
  • Speed research; increase productivity; generate more consistent results; lower analysis costs; and better utilize student efforts.
  • Improve success rates of multiplex assays.
• Diagnostic laboratories, genetic analysis service and CRO organizations can:
  • Lower costs and barriers, eliminate mistakes, and simplify laboratory processes.

Celadon Mod-NA (Modified Nucleic Acids analysis)

Celadon and its partners have developed laboratory and statistical methods and computer code that are generally applicable to the analysis of any modified chemistry. In recent years, research has shown that such modified nucleic acid chemistries can overcome the hybridization limitations of DNA and RNA.

To explain further, genomes are becoming heavily annotated with important features that are key to the next level of research and the creation of medical diagnostics. Analysis of these features often employs oligonucleotides that hybridize at defined locations. When the defined location lies in a poor sequence context, traditional design strategies may fail. Recent research has shown that a new biochemistry tool, Modified Nucleic Acids, can often overcome such limitations. They can improve genetic assay specificity and sensitivity, especially in difficult assay locations, which are frequently encountered when targeting a genomic feature, such as a SNP. Combining these modified chemistries with DNA or RNA is analogous to creating an alloy, a combination with enhanced properties.

Integration of these modified nucleic acid analogues into oligonucleotides, and the application of accurate biophysical design rules for this integration, improves the specificity and stability of oligonucleotides. This is especially true in problematic sequence contexts such as AT or GC rich sequences, sequence locations that have inherent secondary structures such as hairpins and bi-molecular hybridizations, and gene sequences that are repeated in the genome due to descent from a common ancestor.

The Celadon Mod-NA technology gives Celadon the ability to quickly position itself with advantage, or so enable its partner(s), as other modified chemistry methods advance into the market.

Applications: Research, Diagnostics, Multiplex, Pathogen Identification, Personal Genomics, Companion Diagnostics

Initial Methods: LNA Hybridized to DNA, Primers and Probes

Future Methods: LNA Hybridized to RNA, Morpholinos, Zip Nucleic Acid (ZNA), Nucleic Acid Nanotechnology, Peptide Nucleic Acid (PNA), iso-C, iso-G

First Application of Celadon Mod-NA: Celadon LNA Rules

A set of biophysical databases and unique understanding derived in the context of laboratory investigations conducted by Celadon scientists using its Mod-NA analysis methods and analysis has resulted in a ProbITy-compatible technology described herein as the separate product, Celadon LNA Rules.

Future Applications of Celadon Mod-NA

• Custom oligonucleotide synthesis companies can increase sales of high-profit specialty chemicals.
• Point-of-care and other diagnostic device or assay developers can develop research and diagnostic assays faster, with less optimization and cost, while saving component cost due to optimal design rules that result in least number of expensive modified bases or other specialty chemicals. Note that the increased cost of chemistry is trivial compared to total cost of kit.
• Developers of pre-designed research assays can develop assays faster, with less optimization and cost.
• Large pharmaceutical, biotech, and other life science institutions can improve consistency and quality, lower the cost of research and development by increasing assay design success rate, signal strength, and signal consistency.
• Academic and industry researchers can speed research; increase productivity; generate more consistent results; lower analysis costs; and better utilize student efforts.
• Diagnostic laboratories, genetic analysis service and CRO organizations can lower costs and barriers, eliminate mistakes, and simplify laboratory processes.