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公开(公告)号：US09169512B2
公开(公告)日：2015-10-27
申请号：US13380018
申请日：2010-07-01
申请人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Michael M. Becker , Norman C. Nelson , Lyle J. Arnold, Jr.
发明人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Michael M. Becker , Norman C. Nelson , Lyle J. Arnold, Jr.
CPC分类号： C12Q1/6865 , C12Q1/6844 , C12Q1/6848 , C12Q1/6853 , C12Q2549/119 , C12Q2537/143 , C12Q2531/143
摘要： Compositions, reaction mixtures, and methods for performing an amplification reaction, including multiplex amplification reaction, wherein the method comprises using one or more amplification oligomer complexes comprising linked first and second amplification oligomer members. In one aspect, the amplification oligomer complex is hybridized to a target nucleic acid, the target nucleic acid with hybridized amplification oligomer complex is then captured, and other components are washed away. Target sequences of the target nucleic acids are pre-amplified to generate a first amplification product. The first amplification product is amplified in one or more secondary amplification reactions to generate second amplification products.
摘要翻译： 用于进行扩增反应的组合物，反应混合物和方法，包括多重扩增反应，其中所述方法包括使用包含连接的第一和第二扩增寡聚体成员的一种或多种扩增寡聚体复合物。 一方面，将扩增寡聚物复合物与靶核酸杂交，然后捕获具有杂交扩增寡聚物复合物的靶核酸，并洗涤其它组分。 靶核酸的靶序列被预扩增以产生第一扩增产物。 第一扩增产物在一个或多个二级扩增反应中扩增以产生第二扩增产物。
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公开(公告)号：US08642268B2
公开(公告)日：2014-02-04
申请号：US13460341
申请日：2012-04-30
申请人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Norman C. Nelson , Lyle J. Arnold, Jr. , Michael M. Becker
发明人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Norman C. Nelson , Lyle J. Arnold, Jr. , Michael M. Becker
CPC分类号： C12Q1/6881 , C12Q1/6834 , C12Q1/6844 , C12Q1/6865 , C12Q2525/155 , C12Q2525/186 , C12Q2525/197 , C12Q2531/143 , C12Q2565/543
摘要： Compositions that are used in nucleic acid amplification in vitro are disclosed, which include a target specific universal (TSU) promoter primer or promoter provider oligonucleotide that includes a target specific (TS) sequence that hybridizes specifically to a target sequence that is amplified and a universal (U) sequence that is introduced into the sequence that is amplified, by using a primer for the universal sequence. Methods of nucleic acid amplification in vitro are disclosed that use one or more TSU oligonucleotides to attached a U sequence to a target nucleic acid in a target capture step and then use a primer for a U sequence in subsequent amplification steps performed in substantially isothermal conditions to make amplification products that contain a U sequence that indicates the presence of the target nucleic acid in a sample.
摘要翻译： 公开了用于体外核酸扩增的组合物，其包括目标特异性通用（TSU）启动子引物或启动子提供者寡核苷酸，其包括与扩增的靶序列特异性杂交的靶特异性（TS）序列和通用 （U）序列，通过使用通用序列的引物引入扩增的序列。 公开了体外核酸扩增方法，其中使用一种或多种TSU寡核苷酸将U序列连接到目标捕获步骤中的靶核酸，然后在基本上等温条件下进行的随后扩增步骤中使用U序列引物 制备含有指示样品中靶核酸存在的U序列的扩增产物。
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公开(公告)号：US5639604A
公开(公告)日：1997-06-17
申请号：US161706
申请日：1993-12-03
IPC分类号： C12Q1/68 , G01N33/53 , G01N33/532 , G01N33/533 , G01N33/542 , G01N33/58 , C07H21/04 , C12Q1/00
CPC分类号： G01N33/58 , C12Q1/6816 , C12Q1/6823 , G01N33/5306 , G01N33/532 , G01N33/533 , G01N33/542
摘要： Improved homogenous diagnostic assay methods and labels for detecting an analyte in a medium when the analyte is a member of a specific binding pair. The methods and labels provide procedures for reducing background and increasing sensitivity. The binding partner of the analyte is labeled with a substance, the stability of which detectably changes whenever said analyte is bound as a member of the specific binding pair. In a closely related system, the analyte is labeled with a substance susceptible to differential degradation depending on whether or not the analyte is bound as a member of its specific binding pair. After incubation and selective degradation or chemical or biochemical alteration, the amount of analyte bound is detected by measuring either the stability change or the extent of degradation of the label. In a particular system, chemiluminescent acridinium ester labeled probes are used in a homogenous hybridization assay format for sensitively detecting the presence of complement any target polynucleotide sequences.
摘要翻译： 当分析物是特异性结合对的成员时，改进的同源诊断测定方法和用于检测培养基中分析物的标记。 方法和标签提供了降低背景和提高灵敏度的程序。 分析物的结合配偶体用物质标记，当所述分析物作为特异性结合对的成员结合时，其稳定性可检测地改变。 在紧密相关的系统中，根据分析物是否作为其特异性结合对的成员结合，分析物被易受差异降解敏感的物质标记。 孵育和选择性降解或化学或生物化学改变后，通过测量标签的稳定性变化或降解程度来检测分析物的结合量。 在特定系统中，化学发光的吖啶酯标记的探针以均质杂交测定形式使用，以敏感地检测补体存在任何靶多核苷酸序列。
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公开(公告)号：US5185439A
公开(公告)日：1993-02-09
申请号：US332939
申请日：1988-12-12
IPC分类号： C07H21/00
CPC分类号： C07H21/00
摘要： Methods for the construction, labelling and subsequent purification of nucleic acid probes containing primary amines with acridinium esters 4-(2-succinimidyloxycarbonyl-ethyl)phenyl-10-methylacridinium-9-carboxylate fluorosulfonate). The method for attaching acridinium esters to probes uses high (0.1 to 50 mM) acridinium ester concentrations achieved using organic solvent in concentrations of 20% to 80% by volume, and may be carried out either in solution, or with one or the other of the acridinium ester or the probe suspended in solution. Purification (the separation of labelled probe from unlabelled probe and free label) involves (1) first removing most of the free acridinium ester label from probe using rapid separation techniques, then (2) removing substantially all remaining free label from the probe and separating labelled probe from unlabelled probe, involves specific applications of ion exchange, reverse phase or hydroxyapatite HPLC.
摘要翻译： 含有伯胺与吖啶酯4-（2-琥珀酰亚氨基氧羰基 - 乙基）苯基-10-甲基吖啶-9-甲酸氟代磺酸酯的核酸探针的构建，标记和随后纯化的方法。 将吖啶酯连接到探针的方法使用浓度为20％至80％（体积）的有机溶剂获得的高（0.1至50mM）吖啶酯浓度，并且可以在溶液中或与其中的一种或一种 吖啶酯或探针悬浮在溶液中。 纯化（标记的探针与未标记的探针和游离标记物的分离）包括：（1）首先使用快速分离技术从探针中除去大部分游离的吖啶酯标记物，然后（2）从探针中基本上除去所有剩余的游离标记并分离标记 来自未标记探针的探针涉及离子交换，反相或羟基磷灰石HPLC的具体应用。
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公开(公告)号：US20160017392A1
公开(公告)日：2016-01-21
申请号：US14773362
申请日：2014-03-14
申请人： Lyle J. ARNOLD , Norman C. NELSON
发明人： Lyle J. Arnold , Norman C. Nelson
IPC分类号： C12P19/34
CPC分类号： C12P19/34 , C12Q1/6853 , C12Q1/6865 , C12Q2525/186 , C12Q2533/101 , C12Q2565/515 , C12Q2565/537 , C12Q2531/143
摘要： The present invention provides methods for amplifying a nucleic acid from a sample containing a mixture of nucleic acids utilizing a solid support. Methods are provided utilizing user-defined primer oligonucleotides for directional amplification that assists in further manipulation of the target nucleic acid, such as sequencing. Methods are also provided utilizing blocker and displacer oligonucleotides for generating amplified target nucleic acids of defined length. One of these methods provides a first oligonucleotide and a second oligonucleotide affixed to a solid support or separate solid supports. The first oligonucleotide is blocked to prevent extension from the 3′-terminus and has a sequence complementary to a first portion of a target nucleic acid. The second oligonucleotide has a sequence that is identical to a second portion of the target nucleic acid. In this method, a sample is applied to the solid support and the target nucleic acid within the sample binds said first oligonucleotide. The solid support is then washed to remove unbound nucleic acids. A primer sequence containing a target binding region and a polymerase promoter sequence is then annealed to the bound target nucleic acid and extended producing a first duplex nucleic acid. The target sequence is then removed leaving a first nucleic acid that can now bind the second oligonucleotide. The second oligonucleotide is extended to produce a second duplex nucleic acid that contains a second nucleic acid. The second nucleic acid is then amplified by adding a polymerase.
摘要翻译： 本发明提供了使用固体支持物从含有核酸混合物的样品中扩增核酸的方法。 使用用于定向扩增的用户定义的引物寡核苷酸提供了有助于进一步操纵靶核酸的方法，例如测序。 还提供了利用用于产生具有一定长度的扩增靶核酸的阻断剂和置换寡核苷酸的方法。 这些方法之一提供了固定在固体支持物上或第一个固体支持物上的第一寡核苷酸和第二寡核苷酸。 封闭第一寡核苷酸以防止从3'末端延伸并具有与靶核酸的第一部分互补的序列。 第二寡核苷酸具有与靶核酸的第二部分相同的序列。 在该方法中，将样品施加到固体支持物上，并且样品内的靶核酸结合所述第一寡核苷酸。 然后洗涤固体支持物以除去未结合的核酸。 然后将含有靶结合区和聚合酶启动子序列的引物序列退火至结合的靶核酸，延伸产生第一双链体核酸。 然后除去靶序列，留下现在可以结合第二寡核苷酸的第一核酸。 扩展第二寡核苷酸以产生含有第二核酸的第二双链体核酸。 然后通过加入聚合酶扩增第二个核酸。
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公开(公告)号：US08512955B2
公开(公告)日：2013-08-20
申请号：US12828676
申请日：2010-07-01
申请人： Steven T. Brentano , Dmitry Lyakhov , Norman C. Nelson , James D. Carlson , Michael M. Becker , Lyle J. Arnold, Jr.
发明人： Steven T. Brentano , Dmitry Lyakhov , Norman C. Nelson , James D. Carlson , Michael M. Becker , Lyle J. Arnold, Jr.
CPC分类号： C12Q1/6865 , C12Q1/6844 , C12Q1/6848 , C12Q1/6853 , C12Q2549/119 , C12Q2537/143 , C12Q2531/143
摘要： Compositions, reaction mixtures, and methods for performing an amplification reaction, including multiplex amplification reaction, wherein the method comprises using one or more amplification oligomer complexes comprising linked first and second amplification oligomer members. In one aspect, the amplification oligomer complex is hybridized to a target nucleic acid, the target nucleic acid with hybridized amplification oligomer complex is then captured, and other components are washed away. Target sequences of the target nucleic acids are pre-amplified to generate a first amplification product. The first amplification product is amplified in one or more secondary amplification reactions to generate second amplification products.
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公开(公告)号：US20120178636A1
公开(公告)日：2012-07-12
申请号：US13380018
申请日：2010-07-01
申请人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Michael M. Becker , Norman C. Nelson , Lyle J. Arnold, JR.
发明人： Steven T. Brentano , Dmitry Lyakhov , James D. Carlson , Michael M. Becker , Norman C. Nelson , Lyle J. Arnold, JR.
IPC分类号： C40B30/00
CPC分类号： C12Q1/6865 , C12Q1/6844 , C12Q1/6848 , C12Q1/6853 , C12Q2549/119 , C12Q2537/143 , C12Q2531/143
摘要： Compositions, reaction mixtures, and methods for performing an amplification reaction, including multiplex amplification reaction, wherein the method comprises using one or more amplification oligomer complexes comprising linked first and second amplification oligomer members. In one aspect, the amplification oligomer complex is hybridized to a target nucleic acid, the target nucleic acid with hybridized amplification oligomer complex is then captured, and other components are washed away. Target sequences of the target nucleic acids are pre-amplified to generate a first amplification product. The first amplification product is amplified in one or more secondary amplification reactions to generate second amplification products.
摘要翻译： 用于进行扩增反应的组合物，反应混合物和方法，包括多重扩增反应，其中所述方法包括使用包含连接的第一和第二扩增寡聚体成员的一种或多种扩增寡聚体复合物。 一方面，将扩增寡聚物复合物与靶核酸杂交，然后捕获具有杂交扩增寡聚物复合物的靶核酸，并洗涤其它组分。 靶核酸的靶序列被预扩增以产生第一扩增产物。 第一扩增产物在一个或多个二级扩增反应中扩增以产生第二扩增产物。
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8.发明授权Polycationic supports and nucleic acid purification separation and hybridization 失效用于核酸纯化，分离和杂交的聚阳离子载体
公开(公告)号：US5599667A
公开(公告)日：1997-02-04
申请号：US311289
申请日：1994-09-23
CPC分类号： C07H21/00 , C07H1/06 , C12Q1/6806 , C12Q1/6834 , C12Q1/689
摘要： Described herein is the use of polycationic solid supports in the purification of nucleic acids from solutions containing contaminants. The nucleic acids non-covalently bind to the support without significant binding of contaminants permitting their separation from the contaminants. The bound nucleic acids can be recovered from the support. Also described is the use of the supports as a means to separate polynucleotides and hybrids thereof with a nucleotide probe from unhybridized probe. Assays for target nucleotide sequences are described which employ this separation procedure.
摘要翻译： 本文描述的是使用聚阳离子固体载体从含有污染物的溶液中纯化核酸。 核酸与载体非共价结合，而不会使污染物与污染物分离而明显结合污染物。 结合的核酸可以从载体中回收。 还描述了使用载体作为用来自未杂交的探针的核苷酸探针分离多核苷酸和其杂交体的手段。 描述了使用这种分离方法的靶核苷酸序列的测定。
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公开(公告)号：US5451503A
公开(公告)日：1995-09-19
申请号：US255553
申请日：1994-06-07
CPC分类号： C12N15/1068 , C12Q1/6816 , C12Q1/6823 , Y10T436/143333
摘要： Nucleic acid hybridization probes having at least one nucleic acid strand which has at least two separate target specific regions that hybridize to a target nucleic acid sequence, and at least two distinct arm regions that do not hybridize with the target nucleic acid but possess complementary regions that are capable of hybridizing with one another. These regions are designed such that, under appropriate hybridization conditions, the complementary arm regions will not hybridize to one another in the absence of the target nucleic acid; but, in the presence of target nucleic acid the target-specific regions of the probe will anneal to the target nucleic acid, and the complementary arm regions will anneal to one another, thereby forming a branched nucleic acid structure.
摘要翻译： 具有至少一个核酸链的核酸杂交探针，其具有与靶核酸序列杂交的至少两个分离的靶特异性区域，以及至少两个不与靶核酸杂交但具有互补区域的不同臂区域， 能够彼此杂交。 这些区域被设计成使得在适当的杂交条件下，互补臂区不会在不存在靶核酸的情况下彼此杂交; 但是在目标核酸的存在下，探针的靶特异性区域将与靶核酸退火，并且互补臂区域将彼此退火，由此形成支化核酸结构。
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公开(公告)号：US10590453B2
公开(公告)日：2020-03-17
申请号：US15998679
申请日：2018-08-16
申请人： Lyle J. Arnold , Norman C. Nelson
发明人： Lyle J. Arnold , Norman C. Nelson
IPC分类号： C12P19/34 , C12Q1/6844
摘要： The present invention provides methods of amplifying a target nucleic acid utilizing a circularized template. Circularization may be achieved utilizing a bridging oligonucleotide or an inverter primer. The bridging oligonucleotide or inverted primer is extended forming a concatemeric amplicon that can then be used as a template to provide exponential amplification of the target nucleic acid.
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