SEQUENCING VIA ION-TRAP/TIME-OF-FLIGHT MASS SPECTROMETRY

通过离子阱/飞行时间质谱进行测序

• 批准号: 2208964
• 负责人: David M. Lubman
• 金额: $ 13.39万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-09-01 至 1995-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2208964
• 关键词: DNA; biomedical equipment development; computer simulation; mass spectrometry; nucleic acid sequence

We propose to develop new methodology for the rapid sequencing of DNA, involving a combination of biochemical methods and mass spectroscopic analysis. This will involve using mass analysis to sequence the DNA fragments produced by the Sanger method rather than gel electrophoretic techniques. The sequence is encoded in the mass of the DNA strands produced, which can be rapidly separated and identified based upon electrostatic acceleration in a time-of-flight device. Using this methodology it is estimated that at least 50 K bases can be sequenced per day with a desired ultimate goal of 200 K bases per day. This sequencing rate will be based upon matrix-assisted laser desorption/ionization (MALDI) of DNA strands of at least 200 base pairs which can be mass resolved and identified and a multisample desorption probe which will provide rapid sample throughout. In order to enhance the sensitivity of the method to detect the low femtomolar levels of each DNA strand available from biochemical degradation, ion trap storage technology will be interfaced to a reflection time-of-flight device. In addition, the ion trap storage method will be essential for attaining sufficient resolution for the highly energetic ions produced by the MALDI process. Ultimately, a key issue to be explored in this work will involve the development of methods for volatilization and ionization of DNA using MALDI from liquid or solid doped matrices which will be studied and optimized for these experiments.

我们建议开发新的方法来快速测序DNA，