Simultaneous detection of multiple mutations conferring streptomycin resistance inMycobacterium tuberculosis using nanoscale engineered biomagnetites
Simultaneous detection of multiple mutations conferring streptomycin resistance inMycobacterium tuberculosis using nanoscale engineered biomagnetites
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使用纳米级工程生物磁铁矿同时检测结核分枝杆菌中链霉素耐药性的多个突变
DOI：
10.1007/bf02697261
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
T. Matsunaga
中科院分区：
文献类型：
--
作者：
Kohei Maruyama;Norikuni Uchida;H. Takeyama;Tetsushi Mori;R. Kawaguchi;T. Matsunaga
结核分枝杆菌的链霉素耐药主要是由两类不同的突变引起的，包括therpsL基因（3个突变位点）和therrs基因（7个突变位点）内的点突变。我们建立了一种基于热解离曲线分析的链霉素耐药M多突变自动化同时检测系统。使用链霉亲和素标记的细菌磁性颗粒（SA-BacMP）检测结核病。考虑到时间和成本效益，我们使用较少的PCR反应，使用包括多个突变位点的长PCR靶标（rpsL，182 bp;rrs，467 bp）。为了提高通过链霉抗生物素蛋白-生物素结合捕获在BacMP上的靶DNA的量，检查了几种反应条件，例如缓冲液中的盐作为盐种类和浓度，以及反应温度。与常用的1 M NaCl溶液相比，在2 M LiCl溶液中，SA-BacMP上捕获的DNA的量约为6倍（约5 pmol/50 μg BacMP）。在此条件下，成功地同时对链霉素耐药菌株和野生型菌株的rpsL和rrs基因中的10个靶基因进行了自动化核苷酸鉴别。
Streptomycin-resistantMycobacterium tuberculosis has been attributed to two distinct classes of mutations, including point mutations within therpsL gene (three mutation sites) and therrs gene (seven mutation sites). We have developed an automated simultaneous detection system of multiple mutations based on thermal dissociation curve analysis for streptomycin resistance inM. tuberculosis using streptavidin-labeled bacterial magnetic particles (SA-BacMPs). With consideration for time and cost effectiveness, we used fewer PCR reactions, with a long PCR target (rpsL, 182 bp;rrs, 467 bp) including multiple mutation sites. In order to improve the amount of target DNA captured on BacMPs through streptavidin-biotin binding, several reaction conditions, such as salt species and concentration in the buffer, and reaction temperature were examined. Compared to the commonly used 1M NaCl solution, the amount of DNA captured on SA-BacMPs was about six times greater (approx 5 pmoles/50 μg BacMPs) in the 2M LiCl solution. Under these conditions, automated nucleotide discriminations of 10 targets inrpsL andrrs genes of streptomycin-resistant and wild-type strains were successfully performed at the same time.
影响因子：
7.4
作者：
X. Tong;L. Smith
通讯作者：
X. Tong;L. Smith
影响因子：
15
作者：
Hribar, B;Southall, NT;Dill, KA
通讯作者：
Dill, KA
DOI：
10.1054/tuld.1998.0002
发表时间：
1998-01-01
期刊：
Tubercle and Lung Disease
影响因子：
--
作者：
Ramaswamy, S.;Musser, James M.
通讯作者：
Musser, James M.