Visualization ofanoriCregion onanisolated single whole-genome DNA withtriplex forming PNAprobeusing fluorescence microscopy
Visualization ofanoriCregion onanisolated single whole-genome DNA withtriplex forming PNAprobeusing fluorescence microscopy
复制标题
使用荧光显微镜对具有三链体形成 PNA 探针的孤立的单个全基因组 DNA 上的 anoriC 区域进行可视化
DOI：
--
复制
发表时间：
2007
期刊：
影响因子：
--
通讯作者：
T. Imanaka
中科院分区：
文献类型：
--
作者：
Y. Mori;H. Oana;H. Atomi;T. Imanaka
个体mb大小的基因组DNA的伸长。本文提出了一种分析完整报告的方法，将热嗜古菌的细胞，温度大小的dna（可获得柯达卡拉球菌KODI[6]的空间信息）在单分子水平上对特定序列的位置进行了检测。我们利用qdot -链霉亲和素缀合物的复合物，并使用光学镊子进行转移。光学镊子生物素化PNA形成一个三联形荧光探针和一个与实验相比介电常数更大的物体，以观察由于复制（或复制）嗜热古细菌柯达卡拉热球菌（Thermococcus kodakaraensis KODL）完整的基因组DNA而被捕获的染色体圆形溶液和细胞膜的起源位点区域。它的大介电常数。由于KODi基因组DNA沿着DNA部分地提高了空间分辨率，因此DNA被附着在破裂细胞的破坏细胞膜上，在溶液中被光学镊子拖拽和拉长。dna可以通过在膜的kodi残骸的ori区域捕获与同嘌呤序列结合的pnabi被拖到溶液中。当基因组DNA通过凝胶电泳迁移转移试验转移到高盐浓度区域时，证明了单分子水平DNA的荧光光学定位表现出从紧凑到构象转变。然而，由于环境完全消除变化的困难，我们还没有将理论状态检测为随机缠绕状态，并伴有特定区域。通过诱捕和拖拽非特异性结合在qdot -链亲和素之间
elongation ofindividual Mb-size genomic DNA [5]. Inthe Thispaperpresents a methodology foranalyzing intactreport, cells ofthehyperthemophilic archaeon, ThermoMb-size DNAwhichenables toobtain spatial information on coccus kodakaraensis KODI[6]weresubjected toosmotic theposition ofthespecific sequence insingle molecular level.shockandthegenomic DNA released fromtheburst cells We utilized thecomplex ofQdot-streptavidin conjugate and wastransferred using optical tweezers. Theoptical tweezers biotinylated PNA asatriplex forming fluorescent probeand trapan object oflarger permittivity compared tothesurtried tovisualize a region oftheorigin siteofchromosomal rounding solution andcell membrane canbetrapped dueto replication (oriC) onanintact genomic DNA ofahyperthermophilic archaeon, Thermococcus kodakaraensis KODL.To itslarge permittivity. Since KODi genomic DNA ispartly improve thespatial resolution along theDNA,theDNAwas attached tothewreckofacell membraneofaruptured cell, dragged andelongated inthesolution bytheoptical tweezers.theDNA canbedragged inthesolution bytrapping the PNAbinding tohomopurine sequence inoriCregion ofKODI wreckofthemembrane. Whenthegenomic DNA was wasconfirmed bythegelelectrophoresis mobility shift assay transferred toaregion ofahighsalt concentration, the andfluorescence optical mapping insingle molecular level DNA exhibited aconformational transition fromacompact wasdemonstrated. However, wehavenotyetdetected theoriC state intoanrandomcoiled state, accompanied bythe region specifically duetothedifficulty ofthecomplete elimi- change initsenvironment. Bytrapping anddragging the nation ofthenon-specific binding betweenQdot-streptavidin