Sequence information can be obtained from single DNA molecules
Sequence information can be obtained from single DNA molecules
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DOI：
10.1073/pnas.0230489100
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发表时间：
2003-04-01
影响因子：
11.1
通讯作者：
Quake, SR
中科院分区：
文献类型：
--
作者：
Braslavsky, I;Hebert, B;Quake, SR
人类基因组草图的完成花费了数年时间，而这只是一个需要来自许多个人和物种的大量 DNA 和 RNA 序列信息的时期的开始。传统测序技术在成本、速度和灵敏度方面存在局限性，导致对测序信息的需求远远超出了现有的能力。已经有一些建议通过开发单个 DNA 分子测序的能力来解决这些问题，但都没有经过实验证明。在此，我们报道了使用 DNA 聚合酶通过荧光显微镜从单个 DNA 分子中获取序列信息。我们以单碱基分辨率监测荧光标记的核苷酸重复掺入单个 DNA 链，从而可以确定长度达 5 个字节的序列指纹。这些实验表明，可以在单分子水平上研究DNA聚合酶的活性，具有单碱基分辨率和高度并行化，从而为实用的单分子测序技术提供了基础。
The completion of the human genome draft has taken several years and is only the beginning of a period in which large amounts of DNA and RNA sequence information will be required from many individuals and species. Conventional sequencing technology has limitations in cost, speed, and sensitivity, with the result that the demand for sequence information far outstrips current capacity. There have been several proposals to address these issues by developing the ability to sequence single DNA molecules, but none have been experimentally demonstrated. Here we report the use of DNA polymerase to obtain sequence information from single DNA molecules by using fluorescence microscopy. We monitored repeated incorporation of fluorescently labeled nucleotides into individual DNA strands with single base resolution, allowing the determination of sequence fingerprints up to 5 by in length. These experiments show that one can study the activity of DNA polymerase at the single molecule level with single base resolution and a high degree of parallelization, thus providing the foundation for a practical single molecule sequencing technology.