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公开(公告)号：US5981235A
公开(公告)日：1999-11-09
申请号：US681922
申请日：1996-07-29
申请人： John Shultz , Craig E. Smith , Douglas R. Storts , Paula Brisco , Judy Frederiksen , Susanne Selman , Josephine Grosch
发明人： John Shultz , Craig E. Smith , Douglas R. Storts , Paula Brisco , Judy Frederiksen , Susanne Selman , Josephine Grosch
CPC分类号： C12N15/1013 , C12N15/1003 , C12N15/1006
摘要： Solutions containing nucleic acids are treated with an alkaline protease to digest proteins such as nucleases that degrade the nucleic acids. In the isolation of nucleic acids, a biological sample containing nucleic acids is suspended in a solution containing water, buffer and chelating agent, the pH of the solution is adjusted to at least about 10 by adding a solution of sodium hydroxide and anionic detergent, an alkaline protease is incubated in the solution until nucleases are degraded, the pH of the solution is lowered to reduce activity of the alkaline protease by adding a solution having a pH between 3.5 and 4.5 and the alkaline protease is heat inactivated. Lowering of the pH may produce a cloudy solution which is cleared by centrifuging. Nucleic acids are isolated from the cleared solution by alcohol precipitation, or by using paramagnetic particles or a resin matrix containing silica particles. A chaotropic salt can be used to reversibly bind DNA to the resin matrix.
摘要翻译： 含有核酸的溶液用碱性蛋白酶处理以消化降解核酸的蛋白质如核酸酶。 在分离核酸时，将含有核酸的生物样品悬浮于含有水，缓冲剂和螯合剂的溶液中，通过加入氢氧化钠和阴离子洗涤剂溶液将溶液的pH调节至至少约10， 将碱性蛋白酶在溶液中孵育直到核酸酶降解，通过加入pH在3.5和4.5之间的碱性蛋白酶被热灭活，溶液的pH被降低以降低碱性蛋白酶的活性。 降低pH可能产生混浊溶液，通过离心清除。 通过醇沉淀或通过使用顺磁性颗粒或含有二氧化硅颗粒的树脂基质从澄清的溶液中分离出核酸。 离液盐可用于将DNA可逆地结合到树脂基质上。
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公开(公告)号：US5523206A
公开(公告)日：1996-06-04
申请号：US217013
申请日：1994-03-23
CPC分类号： C12Q1/6869
摘要： A new process and kit are described that combines methods for generating the nucleotide base sequence of a DNA molecule with an ultra-sensitive silver staining protocol. This new combination of technologies allows for a direct, non-instrument based visualization of electrophoretically separated sequencing fragments. This non-radioactive system includes sequencing the DNA molecule by forming a set of fragments using an enzymatic dideoxy-mediated chain termination method, electrophoretically separating the DNA fragments on a gel medium, and exposing the gel medium to ultra-sensitive silver-staining solutions for a time determined by viewing the silver stain reacted primer extension products.
摘要翻译： 描述了一种新的方法和试剂盒，其结合了用于产生DNA分子的核苷酸碱基序列与超敏感银染色方案的方法。 这种新的技术组合允许直接的，基于非仪器的电泳分离的测序片段的可视化。 该非放射性系统包括通过使用酶双脱氧介导的链终止法形成一组片段来测序DNA分子，将凝胶介质上的DNA片段电泳分离，并将凝胶介质暴露于超敏感银染色溶液 通过观察银染反应引物延伸产物确定的时间。
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3.发明授权
公开(公告)号：US6077664A
公开(公告)日：2000-06-20
申请号：US656664
申请日：1996-05-31
申请人： Michael R. Slater , Fen Huang , James R. Hartnett , Elena Bolchakova , Douglas R. Storts , Paul Otto , Katharine M. Miller , Alexander Novikov , Galina A. Velikodvorskaya
发明人： Michael R. Slater , Fen Huang , James R. Hartnett , Elena Bolchakova , Douglas R. Storts , Paul Otto , Katharine M. Miller , Alexander Novikov , Galina A. Velikodvorskaya
CPC分类号： C12N9/1252 , C12Q1/686 , C12Q1/6869
摘要： The present invention relates to compositions of thermostable DNA polymerases derived from the hyperthermophilic eubacteria. In particular, the present invention comprises thermostable DNA polymerases from the hyperthermophilic eubacterium known as Thermotoga neapolitana. The present invention provides methods for utilizing naturally-occurring and non-naturally-occurring forms of T. neopolitana DNA polymerase. The T. neopolitana DNA polymerases of the present invention are used in combination with other compounds, including but not limited to pyrophosphatase and DNA polymerases from other thermophilic or hyperthermophilic organisms.
摘要翻译： 本发明涉及源于超嗜热真细菌的热稳定DNA聚合酶的组合物。 特别地，本发明包括来自称为Thermotoga neapolitana的超嗜热真细菌的热稳定DNA聚合酶。 本发明提供了利用天然存在和非天然存在形式的新孢子虫DNA聚合酶的方法。 本发明的新孢杆菌DNA聚合酶与其它化合物组合使用，包括但不限于焦磷酸酶和来自其他嗜热或超嗜热生物的DNA聚合酶。
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公开(公告)号：US5654149A
公开(公告)日：1997-08-05
申请号：US472556
申请日：1995-06-07
CPC分类号： C12Q1/6869
摘要： A new process and kit are described that combines methods for generating the nucleotide base sequence of a DNA molecule with an ultra-sensitive silver staining protocol. This new combination of technologies allows for a direct, non-instrument based visualization of electrophoretically separated sequencing fragments. This non-radioactive system includes sequencing the DNA molecule by forming a set of fragments using an enzymatic dideoxy-mediated chain termination method, electrophoretically separating the DNA fragments on a gel medium, and exposing the gel medium to ultra-sensitive silver-staining solutions for a time determined by viewing the silver stain reacted primer extension products.
摘要翻译： 描述了一种新的方法和试剂盒，其结合了用于产生DNA分子的核苷酸碱基序列与超敏感银染色方案的方法。 这种新的技术组合允许直接的，基于非仪器的电泳分离的测序片段的可视化。 该非放射性系统包括通过使用酶双脱氧介导的链终止法形成一组片段来测序DNA分子，将凝胶介质上的DNA片段电泳分离，并将凝胶介质暴露于超敏感银染色溶液 通过观察银染反应引物延伸产物确定的时间。
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