Rapid PCR of Bacillus anthracis DNA

描述（由申请人提供）：炭疽或毛索氏病是由炭疽芽孢杆菌引起的人畜共患传染病。在“2001年炭疽信件恐慌”的高峰期，美国陆军传染病医学研究所每天检测2000多个样本。目前，USAMRIID几乎所有的炭疽检测都依赖于微生物学方法，这需要一到两天的时间来完成。使用基于聚合酶链反应的方法可以更快、更全面地进行诊断；特别是使用为高速扩增100到500个碱基对长的DNA片段而设计的新机器。迄今为止使用的最快的PCR机是带有数字可编程电子阀的加压He/CO2气体热循环器。利用该气体爆发装置，在不到4分钟的时间内，通过30个PCR循环，将炭疽芽胞杆菌（疫苗）菌株368 bp的DNA片段扩增108倍。使用更短的（151 -288 b.p）扩增和/或更少的（20-25）PCR周期，可以更快地（1.5至3.0分钟）扩增炭疽杆菌质粒编码的毒力因子（pag, left, cya, capA, capC）。目前的实验旨在提高这种高性能He/CO2气体热循环器的易用性，以便于明年在USAMRIID部署。

DESCRIPTION (provided by applicant): Anthrax or Woolsorter's Disease is a zoonotic infection caused by Bacillus anthracis. During the peak of the 'anthrax letter scare of 2001', the US Army Medical Research Institute for Infectious Diseases was testing over 2000 samples per day. At the present time, nearly all anthrax testing at USAMRIID relies on microbiological methods, which require one to two days to carry out. Much faster and more informative diagnosis is possible using PCR-based methods; in particular using new machines designed for high-speed amplification of DNA fragments 100 to 500 base pairs long. The fastest PCR machine employed to date is a pressurized He/CO2 gas thermocycler with digitally programmable electronic valves. Using this gas burst device, a 368 b.p. DNA fragment from B. anthracis Sterne (vaccine) strain was amplified 108-fold through 30 PCR cycles in <4 minutes. Even faster (1.5 to 3.0 minute) PCR-amplification of B. anthracis plasmid-encoded virulence factors (pag, lef, cya, capA, capC) is possible using shorter (151 -288 b.p.) amplicons and/or fewer (20-25) PCR cycles. The present experiments seek to improve the ease-of-use of this high performance He/CO2 gas thermocycler, for deployment at USAMRIID within the next year.