通过DNA结合酶介导的DNA转化为密集功能化的核酸聚合物
Ryan Hili1, Jia Niu, David R Liu
1Howard Hughes Medical Institute, Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts, 02138, United States.
Journal of the American Chemical Society
|December 22, 2012
概括
研究人员使用DNA模板和T4DNA结合酶创造了一种新的合成功能核酸的方法. 这种DNA合成技术允许高效,序列特定的创建长,修饰的核酸聚合物.
科学领域:
- 合成生物学 合成生物学
- 分子生物学分子生物学
- 生物化学 生物化学
背景情况:
- 在各种生物应用中,DNA合成至关重要.
- 目前的方法可能缺乏效率或功能组多样性.
- 开发新的核酸合成策略是一个持续的挑战.
研究的目的:
- 开发一种将DNA序列转化为功能密集的核酸的方法.
- 为了使修改核酸的序列特定聚合.
- 整合合成,选择和再生,用于代的图书馆开发.
主要方法:
- 利用T4DNA结合酶进行5'-酸化三核酸的DNA模板聚合,5'-酸化三核酸.
- 在核酸构建块上结合了各种各样的功能组.
- 用于模板再生的深度通风 (外-) DNA聚合酶使用的原料扩展.
- 在体外翻译,选择和模板再生的综合循环.
主要成果:
- 实现了改性核酸的高效,序列特定的聚合.
- 生成的聚合物至少有50个构建块 (150个核酸) 的长度.
- 通过使用原料扩展证明了模板DNA的成功再生.
- 建立了对图书馆开发和优化的代过程.
结论:
- 开发的方法使高功能核酸的高效合成成为可能.
- 这种方法促进了具有多种化学修饰的序列特定聚合物生成.
- 集成系统支持在体外翻译,选择和模板再生的代循环,以促进图书馆的进化.
相关概念视频
Translesion DNA Polymerases
Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
Nucleic Acid Structure
The pentose sugar in DNA is deoxyribose, while in RNA the pentose sugar is ribose. The difference between the sugars is the presence of the hydroxyl group on the ribose's second carbon and a hydrogen on the deoxyribose's second carbon. The phosphate residue attaches to the hydroxyl group of the 5′ carbon of one sugar and the hydroxyl group of the 3′ carbon of the sugar of the next nucleotide, which forms a 5′ to 3′ phosphodiester linkage.
DNA Structure
DNA has a double-helix structure. The...
DNA Structure
DNA has a double-helix structure. The...
Proofreading
Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore, it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Proofreading
Overview
The Replisome
DNA replication is carried out by a large complex of proteins that act in a coordinated matter to achieve high-fidelity DNA replication. Together this complex is known as the DNA replication machinery or the replisome.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with the...
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with the...
Lagging Strand Synthesis
During replication, the complementary strands in double-stranded DNA are synthesized at different rates. Replication first begins on the leading strand. Replication starts later, occurs more slowly, and proceeds discontinuously on the lagging strand.
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...


