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DEVELOPMENT OF A NEW GENERATION DNA SEQUENCER

新一代 DNA 测序仪的开发

基本信息

批准号：
6160097
负责人：
Shaorong Liu
金额：
$ 46.09万
依托单位：
MOLECULAR DYNAMICS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-06-20 至 2002-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6160097
关键词：
DNA biomedical automation biomedical equipment development capillary electrophoresis confocal scanning microscopy high throughput technology nucleic acid sequence photomultiplier

项目摘要

DNA sequencing using microfabricated devices has a lot of advantages; many of them are originated from a chip injector. To list a few, (1) a very narrow band is injected, (2) a uniform signal intensity profile is obtained, (3) sample volume is reduced to sub-muL and (4) extra sample cleanup is performed. However, there are two major limitations associated with microfabricated devices. (a) limited straight channel length and (b) constrained space for reservoir arrangement. A new generation DNA sequencer based a chip injector coupled with conventional capillaries is proposed. Viewed from the perspective of sequencing on a chip, the proposed scheme eliminates the two limitations. Straight channel length becomes unlimited since a capillary of any length can be coupled to a chip injector and the space for reservoirs becomes unlimited as well since many separate chip injectors can be used to host reservoirs due to the flexibility of the capillaries. Seen from the perspective of sequencing using CGE, the proposed scheme brings all advantages of a chip injector to CGE. The sequencer will consist of a 4-PMT rotary confocal scanner with 8x48-capillary arrays. Linear polyacrylamide will be optimized for sequencing separation on the proposed sequencer. Small volume sample preparation will be incorporated to reduce the sample volume and automated instrumentation will be developed to enhance the sample throughput. Through close collaboration with Genome Sequencing Centers (e.g. Genome Therapeutics Corporation), the sequencer will be developed to meet the needs of genome sequencing world. The new generation sequencer will have specifications of 24-hour operation with minimum attention, average read-length of 800-1000 bases, a run-to-run time of less than 1 hour, and a sample volume of 0.5 to 1 muL.

使用微加工设备进行DNA测序具有许多优点;其中许多优点源自芯片注射器。列举几个，（1）注入非常窄的条带，（2）获得均匀的信号强度分布，（3）样品体积减少到亚μ L，以及（4）进行额外的样品净化。然而，有两个主要的限制与微制造设备。 (a)有限的直通道长度和（B）用于储存器布置的受限空间。提出了一种基于芯片进样器和毛细管的新一代DNA测序仪。从芯片上测序的角度来看，所提出的方案消除了这两个限制。直通道长度变得不受限制，因为任何长度的毛细管可以连接到芯片注射器，并且用于储存器的空间也变得不受限制，因为由于毛细管的柔性，许多单独的芯片注射器可以用于容纳储存器。从测序的角度来看，使用CGE，所提出的方案带来了芯片注入CGE的所有优点。测序仪将由具有8x 48毛细管阵列的4-PMT旋转共聚焦扫描仪组成。线性聚丙烯酰胺将在拟定测序仪上优化测序分离。小体积样品制备将被纳入，以减少样品体积和自动化仪器将被开发，以提高样品的吞吐量。通过与基因组测序中心（如Genome Therapeutics Corporation）的密切合作，测序仪将被开发以满足基因组测序世界的需求。新一代测序仪将具有24小时操作的规格，最小关注，平均读取长度为800-1000个碱基，运行到运行时间小于1小时，样品体积为0.5至1穆尔。