Multipurpose reversible terminators

多用途可逆终结器

• 批准号: 8145336
• 负责人: Daniel Hutter
• 金额: $ 64.46万
• 依托单位: FIREBIRD BIOMOLECULAR SCIENCES, LLC
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-17 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8145336
• 关键词: Abbreviations; American; Architecture; Base Sequence; Biological; Biological Assay; Biology; Buffers; C-glycoside; Cicatrix; Cleaved cell; Complex; Contracts; DNA; DNA Sequence; DNA analysis; DNA-Directed DNA Polymerase; Data; Deoxyribonucleotides; Detection; Development; Diagnostic; Disulfides; Emulsions; Enzymes; Excision; Family; Foundations; Funding; Genetic; Genomics; Glycols; Grant; Hereditary Disease; Human; Information Systems; Left; Length; Licensing; Link; Market Research; Medical; Medicine; Modeling; Modification; Molecular Evolution; Mutation; Natural regeneration; Nitrogen; Nitrous Acid; Nucleic Acids; Nucleic acid sequencing; Occupations; Oils; Oligonucleotides; Phase; Polymerase; Polymorphism Analysis; Primer Extension; Production; Pyrimidine; RNA-Directed DNA Polymerase; Reading; Reagent; Recovery; Research; Risk; Sales; Sampling; Science; Scientist; Series; Side; Single Nucleotide Polymorphism; Small Business Technology Transfer Research; Sodium Nitrite; Structure; Sulfhydryl Compounds; Synthesis Chemistry; System; Technology Transfer; Therapeutic; Time; Tissues; Transferase; Water; Work; appendage; base; dideoxynucleotide; effectiveness research; functional group; innovation; metaperiodate; molecular recognition; mutant; next generation; nucleic acid analog; nucleoside triphosphate; phosphoramidite; public health relevance; software development; tool; tripolyphosphate; two-dimensional

DESCRIPTION (provided by applicant): This Phase 2 STTR application will allow the Foundation for Applied Molecular Evolution (FfAME) to complete its transfer of technology to Firebird Biomolecular Sciences LLC, allowing Firebird to commercialize nucleoside triphosphates where the 3'-OH groups are blocked by a 3'-ONH2 group. Triphosphates having this modification are accepted by DNA polymerases and other enzymes. After being incorporated at the 3'-end of a growing oligonucleotide, the 3'-ONH2 group blocks extension (just as a 2',3'-dideoxynucleotide does). However, this blockage is reversible; buffered sodium nitrite rapidly cleaves the O-NH2 bond to generate the natural 3'-OH group, yielding under mild conditions a primer that can again be extended by DNA polymerase. Further, the 3'-ONH2 group offers a handle that allows the terminated DNA to selectively react with certain functional groups, allowing for the recovery of the extended, terminated DNA from complex biological mixtures. This is useful in many architectures that analyze DNA, including "next generation" sequencing that promises to personalize diagnostics and therapeutics, tools to detect single nucleotide polymorphisms, strategies to do "effectiveness research" for medical treatments, and other architectures that have increasing complexity and risk. Therefore, the products to be developed here have multiple applications in a variety of research settings, genetic disease discovery, and human diagnostics. This Phase 2 project, together with Firebird's internal funding, will build a production pipeline to prepare 12 different triphosphates carrying the reversible terminator, as well as four phosphoramidites to support kits that use these. The synthetic chemistry developed in the Phase 1 period will support this pipeline. Further, tools used in the Phase I period to develop polymerases to handle unnatural nucleoside triphosphates will be applied to create still better polymerases for assays that exploit our triphosphates. Finally, kits for DNA analysis will be produced that exploit these reversible terminators, in many cases with other reagent innovations that are proprietary to FfAME, Firebird, and others. Several customers are already exploiting these compounds. These include foreign customers, whose purchase of these reagents is creating American jobs through the sale of products internationally. Growing its portfolio of nucleic acid analogs underlies the next phase in the development of Firebird as a corporation. PUBLIC HEALTH RELEVANCE: Firebird Biomolecular Sciences LLC commercializes innovative reagents, polymerases that exploit them, and assays and architectures based on them, for the detection, analysis and sequencing of nucleic acids. Firebird also develops software to exploit genomic sequence data, and what it means in biology and medicine. This Phase 2 STTR project will create over two dozen jobs through the development of a series of reversible terminators suitable for next generation DNA sequencing, SNP detection, and DNA capture.

描述（由申请人提供）：该第 2 阶段 STTR 申请将允许应用分子进化基金会 (FfAME) 完成向 Firebird Biomolecular Sciences LLC 的技术转让，从而使 Firebird 能够将 3'-OH 基团被封闭的核苷三磷酸商业化由 3'-ONH2 基团组成。具有这种修饰的三磷酸被DNA聚合酶和其他酶所接受。掺入生长寡核苷酸的 3' 端后，3'-ONH2 基团会阻止延伸（就像 2',3'-双脱氧核苷酸一样）。然而，这种阻塞是可逆的；缓冲的亚硝酸钠快速裂解 O-NH2 键，生成天然 3'-OH 基团，在温和条件下产生可再次被 DNA 聚合酶延伸的引物。此外，3'-ONH2 基团提供了一个手柄，允许终止的 DNA 选择性地与某些官能团反应，从而允许从复杂的生物混合物中回收延伸的终止 DNA。这在许多分析 DNA 的架构中都很有用，包括承诺个性化诊断和治疗的“下一代”测序、检测单核苷酸多态性的工具、为医疗治疗进行“有效性研究”的策略，以及其他日益复杂和复杂的架构。风险。因此，这里要开发的产品在各种研究环境、遗传疾病发现和人类诊断中有多种应用。该第二阶段项目与 Firebird 的内部资金一起，将建立一条生产管道，以制备 12 种不同的携带可逆终止子的三磷酸盐，以及四种亚磷酰胺以支持使用这些试剂盒的试剂盒。第一阶段开发的合成化学将支持该管道。此外，第一阶段用于开发处理非天然三磷酸核苷的聚合酶的工具将用于创建更好的聚合酶，用于利用我们的三磷酸的测定。最后，将生产利用这些可逆终止子的 DNA 分析试剂盒，在许多情况下与 FfAME、Firebird 等公司专有的其他试剂创新一起生产。一些客户已经在利用这些化合物。其中包括外国客户，他们购买这些试剂通过在国际上销售产品为美国创造了就业机会。扩大其核酸类似物产品组合是 Firebird 公司下一阶段发展的基础。 公共健康相关性：Firebird Biomolecular Sciences LLC 将创新试剂、利用它们的聚合酶以及基于它们的测定和架构商业化，用于核酸的检测、分析和测序。 Firebird 还开发了软件来利用基因组序列数据及其在生物学和医学中的意义。 STTR 项目第二阶段将通过开发一系列适用于下一代 DNA 测序、SNP 检测和 DNA 捕获的可逆终止子，创造超过 20 个就业岗位。