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31. 发明授权

US5185439A Acridinium ester labelling and purification of nucleotide probes 失效
标题翻译：吖啶酯标记和纯化核苷酸探针
公开(公告)号：US5185439A
公开(公告)日：1993-02-09
申请号：US332939
申请日：1988-12-12
申请人： Lyle J. Arnold, Jr. , Norman C. Nelson
发明人： Lyle J. Arnold, Jr. , Norman C. Nelson
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。

32. 发明申请

US20200002740A9 Methods for amplification of nucleic acids utilizing a circularized template prepared from a target nucleic acid 审中-公开
公开(公告)号：US20200002740A9
公开(公告)日：2020-01-02
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.

33. 发明授权

US10179931B2 Methods for immobilizing target nucleic acids utilizing combinatorial capture probes 有权
公开(公告)号：US10179931B2
公开(公告)日：2019-01-15
申请号：US15652372
申请日：2017-07-18
申请人： Lyle J. Arnold, Jr.
发明人： Lyle J. Arnold, Jr.
IPC分类号： C12P19/34 , C12Q1/6834
摘要： The present invention provides methods for immobilizing target nucleic acids on a solid support utilizing combinatorial capture probe pairs. These pairs contain first and second capture oligonucleotides that each comprise a target binding region, a capture region and a stem region positioned between the target binding and capture regions. The target binding regions comprise nucleic acid sequences that allow them to hybridize to adjacent regions on the target nucleic acid. The stem regions have nucleic acid sequences that are complementary to each other and the capture regions each comprise a sequence that when positioned adjacent to one another produce a combined nucleic acid sequence that is complementary to a portion of an oligonucleotide bound to a solid support. When the first and second capture oligonucleotides are annealed to the target nucleic acid, the stem regions are brought together allowing them to hybridize, which in turn brings the capture regions together to produce a combined nucleic acid sequence. This combined nucleic acid sequence is then able to hybridize to the oligonucleotide bound to the solid support thereby immobilizing the target nucleic acid.

34. 发明申请

US20180010173A1 Methods for Immobilizing Target Nucleic Acids Utilizing Combinatorial Capture Probes 审中-公开
公开(公告)号：US20180010173A1
公开(公告)日：2018-01-11
申请号：US15652372
申请日：2017-07-18
申请人： Lyle J. Arnold
发明人： Lyle J. Arnold
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6834 , C12Q2525/161 , C12Q2525/30 , C12Q2537/162
摘要： The present invention provides methods for immobilizing target nucleic acids on a solid support utilizing combinatorial capture probe pairs. These pairs contain first and second capture oligonucleotides that each comprise a target binding region, a capture region and a stem region positioned between the target binding and capture regions. The target binding regions comprise nucleic acid sequences that allow them to hybridize to adjacent regions on the target nucleic acid. The stem regions have nucleic acid sequences that are complementary to each other and the capture regions each comprise a sequence that when positioned adjacent to one another produce a combined nucleic acid sequence that is complementary to a portion of an oligonucleotide bound to a solid support. When the first and second capture oligonucleotides are annealed to the target nucleic acid, the stem regions are brought together allowing them to hybridize, which in turn brings the capture regions together to produce a combined nucleic acid sequence. This combined nucleic acid sequence is then able to hybridize to the oligonucleotide bound to the solid support thereby immobilizing the target nucleic acid.

35. 发明申请

US20170009282A1 Methods for amplification of nucleic acids utilizing hairpin loop or duplex primers 审中-公开
标题翻译：使用发夹环或双链体引物扩增核酸的方法
公开(公告)号：US20170009282A1
公开(公告)日：2017-01-12
申请号：US14999978
申请日：2016-07-21
申请人： Lyle J. Arnold
发明人： Lyle J. Arnold
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6853 , C12N15/1065 , C12Q2521/501 , C12Q2525/301 , C12Q2563/185
摘要： The present invention provides methods of amplifying a target nucleic acid utilizing duplex primer. The first strand of the primer comprises a random nucleotide sequence of about 6 to about 9 nucleotides in length that is able to hybridize to the target nucleic acid and a tag sequence. The second strand of the primer comprises a sequence complementary to the tag sequence allowing the primer to form a duplex and the ability to bind the tag sequence of the product nucleic acid for further amplification. The resulting nucleic acid produced contains tag sequences on both the 3′- and 5′-termini.
摘要翻译：本发明提供了利用双链体引物扩增靶核酸的方法。引物的第一条链包含长度为约6至约9个核苷酸的随机核苷酸序列，其能够与靶核酸和标签序列杂交。引物的第二条链包含与标签序列互补的序列，使得引物形成双链体并结合产物核酸的标签序列用于进一步扩增的能力。产生的所得核酸含有3'和5'末端的标签序列。

36. 发明申请

US20160068900A1 Methods for Amplifying Fragmented Target Nucleic Acids Utilizing an Assembler Sequence 审中-公开
标题翻译：使用汇编序列扩增分离的靶核酸的方法
公开(公告)号：US20160068900A1
公开(公告)日：2016-03-10
申请号：US14773366
申请日：2014-03-15
申请人： Lyle J. ARNOLD , Norman C. NELSON
发明人： Lyle J. Arnold , Norman C. Nelson
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6858 , C12Q1/6806 , C12Q1/6844 , C12Q2525/161 , C12Q2525/186 , C12Q2525/204 , C12Q2531/101
摘要： The present invention provides methods of amplifying a fragmented target nucleic acid containing short target nucleic acid fragments utilizing an assembler sequence to convert these short fragments into longer sequences enabling their identification and interrogation. This is particularly important when attempting to identify small genetic variations, such as SNVs, present in highly fragmented nucleic acid samples. Amplification is accomplished by hybridizing the short target nucleic acid sequences to the assembler sequence, where these short sequences serve as primers for extension. Since the fragmented target nucleic acids that contain SNVs are utilized as primers on the assembler sequence they are preserved during amplification and can be detected.
摘要翻译：本发明提供了利用汇编序列扩增含有短靶核酸片段的片段化靶核酸的方法，以将这些短片段转化成更长的序列，使其能够进行鉴定和询问。当试图鉴定存在于高度片段化的核酸样品中的小的遗传变异（例如SNV）时，这尤其重要。通过将短目标核酸序列杂交到汇编序列来完成扩增，其中这些短序列用作扩增的引物。由于含有SNV的片段化靶核酸作为汇编序列上的引物，因此在扩增过程中被保留并可以被检测。

37. 发明申请

US20160024546A1 Methods for Amplification of Nucleic Acids Utilizing a Circularized Template Prepared from a Target Nucleic Acid 审中-公开
标题翻译：使用从靶核酸制备的圆形模板扩增核酸的方法
公开(公告)号：US20160024546A1
公开(公告)日：2016-01-28
申请号：US14773363
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 , C12Q1/6844 , C12Q2531/119 , C12Q2531/125 , C12Q2537/162
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.
摘要翻译：本发明提供利用环化模板扩增靶核酸的方法。可以使用桥连寡核苷酸或反向引物实现环化。桥接寡核苷酸或反向引物延伸形成一个可以用作模板以提供靶核酸的指数扩增的并联扩增子。

38. 发明申请

US20110003305A1 METHODS AND COMPOSITIONS FOR NUCLEIC ACID AMPLIFICATION 有权
标题翻译：用于核酸扩增的方法和组合物
公开(公告)号：US20110003305A1
公开(公告)日：2011-01-06
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.
IPC分类号： C12Q1/68 , C12P19/34 , C12N9/00
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。

39. 发明授权

US5585481A Linking reagents for nucleotide probes 失效
标题翻译：连接核苷酸探针的试剂
公开(公告)号：US5585481A
公开(公告)日：1996-12-17
申请号：US182666
申请日：1994-01-14
申请人： Lyle J. Arnold, Jr. , Mark A. Reynolds , Ram S. Bhatt
发明人： Lyle J. Arnold, Jr. , Mark A. Reynolds , Ram S. Bhatt
IPC分类号： C07F9/24 , C07H21/00 , C07H1/02 , C07H19/00
CPC分类号： C07H21/00 , C07F9/2408
摘要： A versatile reagent with a non-nucleotide monomeric unit having a ligand, and first and second coupling groups which are linked to the non-nucleotide monomeric unit. The ligand can be either a chemical moiety, such as a label or intercalator, or a linking arm which can be linked to such a moiety. Such reagent permits preparation of versatile nucleotide/non-nucleotide polymers, having any desired sequence of nucleotide and non-nucleotide monomeric units, each of the latter of which bear a desired ligand. These polymers can for example, be used as probes which can exhibit enhanced sensitivity and/or which are capable of detecting a genus of nucleotides each species of which has a common target nucleotide sequence of interest bridged by different sequences not of interest.
摘要翻译：具有具有配体的非核苷酸单体单元的多功能试剂和与非核苷酸单体单元连接的第一和第二偶联基团。配体可以是化学部分，例如标记或嵌入剂，或可以连接到这种部分的连接臂。这种试剂允许制备通用的核苷酸/非核苷酸聚合物，具有任何所需的核苷酸和非核苷酸单体单元序列，后者各自具有所需的配体。这些聚合物可以例如用作可以显示增强的灵敏度的探针和/或能够检测核苷酸属的核苷酸，其中每个物种具有由不感兴趣的不同序列桥接的共同目标核苷酸序列。

40. 发明授权

US5539097A Oligonucleotide polymeric support system 失效
标题翻译：寡核苷酸聚合物支持体系
公开(公告)号：US5539097A
公开(公告)日：1996-07-23
申请号：US272290
申请日：1994-07-08
申请人： Lyle J. Arnold, Jr.
发明人： Lyle J. Arnold, Jr.
IPC分类号： C07H21/00 , C07H19/067 , C07H19/167 , C07H21/02
CPC分类号： C07H21/00 , Y02P20/55
摘要： A versatile polymeric support system for the synthesis of oligonucleotides is provided featuring a universal primer which allows chain elongation, in either the 3' or 5' direction, with any currently available DNA or RNA synthesis method, by a process which utilizes oxidatively cleaved primers to facilitate chain elongation and release. The support system is capable of withstanding mildly basic and acidic reaction conditions, while still permitting a convenient and quantitative release, either before or after removal of protecting groups from reactive groups, of synthesized oligonucleotides from a single polymeric support. Removal of the protecting groups before cleavage of the oligomer from the support permits the use of the immobilized oligomer as an affinity hybridization support for both isolating and detecting complementary polynucleic acids.
摘要翻译：提供了用于合成寡核苷酸的通用聚合物支持体系，其特征在于通用引物，其通过使用氧化性切割引物的方法，允许以任何目前可用的DNA或RNA合成方法在3'或5'方向上延伸3'或5' 促进链伸长和释放。支持体系能够承受轻度碱性和酸性的反应条件，同时在从单一聚合物载体除去合成的寡核苷酸的保护基团的反应性基团之前或之后仍然允许方便和定量的释放。在从载体裂解低聚物之前除去保护基允许使用固定的寡聚物作为分离和检测互补多核酸的亲和杂交支持物。

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