Evolution of a Deoxyribozyme Ligase

脱氧核酶连接酶的进化

• 批准号: 7151684
• 负责人: Bianca J Lam
• 金额: $ 4.4万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7151684
• 关键词: DNA; biotechnology; brca gene; directed evolution; enzyme activity; enzyme substrate; gene mutation; genetic disorder diagnosis; globin; ligase; nucleic acid amplification techniques; nucleic acid chemical synthesis; phosphoester ligase; point mutation; polymerase chain reaction; postdoctoral investigator; ribozymes; single nucleotide polymorphism; thalassemia

DESCRIPTION (provided by applicant): The potential of DNA as a stable catalyst may give this molecule more success than ribozymes as tools for molecular biology, biotechnology, nanotechnology, and clinical diagnostics. Furthermore, comparison of catalytic RNA structures, which occurs naturally and more frequently, and DNA structures that result in catalytic motifs will prove crucial in revealing fundamental features of biocatalysts. To further gain insight into evolution and the structure and function of nucleic acid enzymes, this proposal will aim to 1) evolve a deoxyribozyme ligase from random sequences using stepwise evolution procedures, 2) optimize a deoxyribozyme-catalyzed ligase chain reaction (LCR), a method for the exponential amplification of target DNA through cycles of denaturing and annealing of ligation substrates and template, and 3) detect genetic mutations using the deoxyribozyme-catalyzed LCR, such as point mutations of BRCA1, BRCA1, and ones that give rise to beta-thalassemia.

描述（由申请人提供）：DNA作为稳定催化剂的潜力可能使这种分子比核酶更成功地作为分子生物学、生物技术、纳米技术和临床诊断的工具。此外，比较催化RNA结构（自然发生且更频繁）和导致催化基序的DNA结构，将证明对揭示生物催化剂的基本特征至关重要。为了进一步深入了解核酸酶的进化以及结构和功能，本项目将致力于：1)利用逐步进化程序从随机序列进化出脱氧核酶连接酶；2)优化脱氧核酶催化的连接酶链反应（LCR），这是一种通过连接底物和模板的变性和退火循环对目标DNA进行指数扩增的方法；3)利用脱氧核酶催化的LCR检测基因突变。比如BRCA1、BRCA1的点突变，以及导致-地中海贫血的点突变。