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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：US4673638A
公开(公告)日：1987-06-16
申请号：US721329
申请日：1985-04-09
IPC分类号： C12Q1/04 , G01N33/569
CPC分类号： G01N33/569 , C12Q1/04
摘要： A method is provided for detecting a microorganism which produces a desired substance. The method involves overlaying a membrane on an agar surface in a predetermined orientation. Microorganisms are grown on the membrane and the substance allowed to pass into the agar. The membrane is removed and a detectable reaction between the substance and one or more reagents is observed. The location of the microorganism to be detected can be determined by the predetermined orientation, and the microorganism can be isolated by removing it from the membrane. When the substance is not secreted by the microorganism, the microorganisms are lysed releasing the substance for passage into the agar. The microorganisms producing the substance can be located on replicate plates and isolated.
摘要翻译： 提供了一种用于检测产生所需物质的微生物的方法。 该方法包括以预定方向将膜覆盖在琼脂表面上。 微生物在膜上生长并允许物质进入琼脂。 去除膜，并观察物质与一种或多种试剂之间的可检测反应。 要检测的微生物的位置可以通过预定取向来确定，并且可以通过将其从膜上除去来分离微生物。 当物质不被微生物分泌时，微生物被裂解释放物质以通入琼脂。 产生物质的微生物可以位于复制板上并分离。
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公开(公告)号：US5543302A
公开(公告)日：1996-08-06
申请号：US159211
申请日：1993-11-30
CPC分类号： C12N9/54 , C11D3/38618 , C12N9/50
摘要： Proteases are susceptible to autolytic degradation. The method of the invention provides a method to identify susceptible autolysis sites which are cleaved to produce a few discrete fragments. Once identified, amino acid(s) at or in the vicinity of the cleavage site may be chemically modified or the DNA sequence capable of encoding the amino acid present at the site may be changed or deleted by site directed mutagenesis to produce an autolysis stable protease. The invention is particularly applicable to alkaline proteases which are produced in large quantities and used as detergent additives. Stable alkaline proteases could then be used in liquid detergent formulations.
摘要翻译： 蛋白酶易于自溶降解。 本发明的方法提供了一种识别敏感的自溶位点的方法，其被切割以产生几个离散的片段。 一旦鉴定，可以化学修饰在切割位点处或其附近的氨基酸，或者可以通过定点诱变来改变或删除存在于位点的氨基酸的DNA序列以产生自溶稳定蛋白酶 。 本发明特别适用于大量生产并用作洗涤剂添加剂的碱性蛋白酶。 稳定的碱性蛋白酶可用于液体洗涤剂配方。
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