ENZYMATIC SYNTHESIS OF DEFINED SEQUENCE DNA

确定序列 DNA 的酶促合成

• 批准号: 2859179
• 负责人: RICHARD H TULLIS
• 金额: $ 9.94万
• 依托单位: SYNGEN RESEARCH
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至 1999-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2859179
• 关键词: enzyme mechanism; method development; nucleic acid chemical synthesis; nucleic acid sequence; nucleotidyltransferase

DESCRIPTION (Adapted from the Investigator's Abstract): Current techniques typically limit synthetic DNA to chain lengths of less than 150 bases and to modifications which can survive strong acid, base, and oxidants. The principal limitations are chemical side-reactions, in particular depurination. An enzymatic approach will allow the synthesis of long oligonucleotides containing chemically sensitive labels. This process is automatable and the unused reagents can be readily recycled. Phase I is designed to determine the feasibility of using polymerases to perform the automated synthesis of defined sequence oligonucleotides. The principal technical innovations in this phase of the proposal involve; 1) chemical synthesis of the nucleotide monomers; 2) synthesis of an appropriate solid phase resin; 3) modification of a DNA synthesizer to perform these syntheses; and 4) enzymatic synthesis of defined sequence oligonucleotides. In Phase II, the synthesis conditions will be refined to allow construction of very long chain oligonucleotides. Nucleotides containing backbone or modified bases will also be synthesized to make oligonucleotides containing chemically sensitive functionalities that are difficult to obtain using conventional means. The first of these products will be marketed during Phase II. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE

描述（改编自研究者摘要）：当前 技术通常将合成DNA的链长限制为小于 150个碱基和修饰，可以生存强酸，碱，和 氧化剂 主要的限制是化学副反应， 特别是脱嘌呤作用。酶促方法将允许合成 含有化学敏感标记的长寡核苷酸。 这 该过程是自动化的，并且未使用的试剂可以容易地再循环。 阶段I被设计为确定使用聚合酶的可行性 以进行确定序列的寡核苷酸的自动化合成。 本阶段的主要技术创新 涉及：1）核苷酸单体的化学合成; 2）合成 合适的固相树脂; 3）DNA的修饰 合成器进行这些合成;和4）酶促合成 确定序列的寡核苷酸。 在第二阶段，合成条件将得到改进， 构建极长链寡核苷酸。 核苷酸 也将合成含有骨架或修饰碱基的化合物， 含有化学敏感官能团的寡核苷酸， 很难用常规方法获得。 其中第一个 产品将在第二阶段上市。 拟议商业应用：不可用