RAPID LOW COST DNA SEQUENCING BY MASS SPECTROMETRY

这项提议的目标是开发一种新型的质谱学 允许500个碱基长(或更长)的测序的方法 DNA链。该仪器的实现将使DNA 以每天接近一百万个碱基的速度进行测序- A的速度比目前可用的技术快三个数量级。要实现 这一目标，我们计划将当前的DNA测序策略与 三项新技术可以生产出低成本、易于自动化的测序 方法。我们打算(I)利用我们在激光汽化方面的专业知识 开发一种新的汽化策略，允许精确控制 解吸的DNA的化学性质，将非常容易迅速 测序；(Ii)利用电离方案，允许选择性地 蒸发DNA链的电离和超灵敏检测 这些链；以及(Iii)对这些电离DNA链进行质量分析 光谱技术将具有所需的灵敏度和分辨率 对长达1000个碱基的DNA进行测序。我们技术的自动化 建议将导致一种极其灵敏的DNA测序方法 需要很少的体力劳动。排序成本是由一个 使用此方法的仪器应低于0.10美元/ 测序的碱基，从而导致至少10倍的 与人类基因组计划相关的测序成本。

The goal of this proposal is the development of a novel mass spectral method which will permit the sequencing of 500 base-long (or longer) strands of DNA. The realization of this instrument will allow DNA sequencing to be performed at a rate approaching one million bases per day- a rate three orders faster than currently available techniques. To achieve this objective, we plan on melding current DNA sequencing strategies with three new technologies to produce a low-cost, easily automated sequencing method. We intend (i) to draw on our expertise in laser vaporization to develop a new vaporization strategy that will permit precise control over the chemical nature of the desorbed DNA and will be very amenable to rapid sequencing; (ii) to utilize an ionization scheme that will allow selective ionization of the vaporized DNA strands and ultrasensitive detection of these strands; and (iii) to analyze these ionized DNA strands by a mass spectral technique that will have the sensitivity and resolution required to sequence DNA up to 1000 bases long. The automation of the technology we propose would result in an extremely sensitive DNA sequencing method requiring little manual labor. The sequencing cost resulting from an instrument that used this methodology should be less than $0.10 per sequenced base, thus resulting in at least a 10-fold reduction in the sequencing costs associated with the Human Genome Initiative.