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1. 发明授权

US06573046B1 Eukaryotic use of improved chimeric mutational vectors 有权
标题翻译：改良的嵌合突变载体的真核生物学应用
公开(公告)号：US06573046B1
公开(公告)日：2003-06-03
申请号：US09429291
申请日：1999-10-28
申请人： Eric B. Kmiec , Howard B. Gamper , Allyson D. Cole-Strauss
发明人： Eric B. Kmiec , Howard B. Gamper , Allyson D. Cole-Strauss
IPC分类号： C12Q168
CPC分类号： C12N15/102
摘要： The invention is based on the reaction of recombinagenic oligonucleotides in a cell-free system containing a cytoplasmic cell extract and a test duplex DNA on a plasmid. The reaction specifically converts a mutant kanr gene to recover the resistant phenotype in transformed MutS, RecA deficient bacteria and allows for the rapid and quantitative comparison of recombinagenic oligonucleobases. Using this system a type of Duplex Mutational Vector termed a Heteroduplex Mutational Vector, was shown to be more active in than the types of mutational vectors heretofore tested. Further improvements in activity were obtained by replacement of a tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2′-O-methyl-uridine, to link the two strands of the Duplex Mutational Vector and removal of the DNA-containing intervening segment. The claims concern Duplex Mutational Vectors that contain the above improvements. In an alternative embodiment the claims concern a reaction mixture containing a recombinagenic oligonucleobase, a cell-free enzyme mixture and a duplex DNA containing a target sequence. In yet an alternative embodiment, the invention concerns the use of such mixture to test improvements in recombinagenic oligonucleobases, as well as to test the effects of compounds on the activity of the cell-free enzyme mixture and also to make specific changes in the target DNA sequence.
摘要翻译：本发明基于重组型寡核苷酸在含有质粒上的细胞质细胞提取物和测试双链体DNA的无细胞系统中的反应。该反应特异性转化突变型kanr基因以在转化的MutS，RecA缺陷型细菌中恢复抗性表型，并允许对重组型寡核苷酸的快速和定量比较。使用该系统，称为异源双链突变载体的一种双链突变载体被证明比迄今为止测试的突变载体的类型更有活性。通过用核酸酶抗性寡核苷酸（例如四 - 2'-O-甲基 - 尿苷）替代四硫氨酸接头，连接双链突变载体的两条链和去除含有DNA的中间片段，获得活性的进一步改善。这些权利要求涉及包含上述改进的双链突变载体。在替代实施方案中，权利要求涉及含有重组型寡核苷酸，无细胞的酶混合物和含有靶序列的双链体DNA的反应混合物。在另一个实施方案中，本发明涉及使用这样的混合物来测试重组型寡核苷酸的改进，以及测试化合物对无细胞酶混合物的活性的影响，并且还对靶DNA进行特异性改变序列。

2. 发明授权

US6010907A Eukaryotic use of non-chimeric mutational vectors 失效
标题翻译：非嵌合突变载体的真核生物学应用
公开(公告)号：US6010907A
公开(公告)日：2000-01-04
申请号：US78064
申请日：1998-05-12
申请人： Eric B. Kmiec , Howard B. Gamper , Allyson D. Cole-Strauss
发明人： Eric B. Kmiec , Howard B. Gamper , Allyson D. Cole-Strauss
IPC分类号： C12N15/09 , C12N1/11 , C12N15/10 , C12N5/10 , C12N15/11 , C12N15/63
CPC分类号： C12N15/102
摘要： The invention is based on the reaction of Duplex Mutational Vector in a cell-free system containing a cytoplasmic cell extract and a test plasmid. The reaction specifically converts a mutant kan.sup.r gene to recover the resistant phenotype in transformed MutS, RecA deficient bacteria. Using this system a type of Duplex Mutational Vector termed a Non-Chimeric Mutational Vector, having no RNA:DNA hybrid-duplex is shown to be an effective substrate for eukaryotic enzymes. The invention concerns the use of Non-Chimeric Mutational Vectors protected from 3' exonuclease attack in eukaryotic cells. Such protection can be conferred by replacement of a tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2'-O-methyl-uridine, to link the two strands of the recombinagenic oligonucleobase.
摘要翻译：本发明基于双链突变载体在含有细胞质细胞提取物和测试质粒的无细胞系统中的反应。该反应特异性转化突变型kanr基因以在转化的MutS，RecA缺陷型细菌中恢复抗性表型。使用该系统，一种称为非嵌合突变载体的双链突变载体，没有RNA：DNA杂合双链体显示为真核生物酶的有效底物。本发明涉及在真核细胞中保护3'核酸外切酶攻击的非嵌合突变载体的用途。这样的保护可以通过用核酸酶抗性寡核苷酸（例如四 - 2'-O-甲基 - 尿苷）替代四连体酶连接体来连接重组型寡核苷酸的两条链。

3. 发明授权

US06936467B2 Targeted chromosomal genomic alterations with modified single stranded oligonucleotides 失效
标题翻译：用修饰的单链寡核苷酸靶向染色体基因组改变
公开(公告)号：US06936467B2
公开(公告)日：2005-08-30
申请号：US09818875
申请日：2001-03-27
申请人： Eric B. Kmiec , Howard B. Gamper , Michael C. Rice
发明人： Eric B. Kmiec , Howard B. Gamper , Michael C. Rice
IPC分类号： A61K38/00 , A61K48/00 , C12N15/10 , C12N15/113 , C12N15/82 , C12N15/09
CPC分类号： C12N15/113 , A61K38/00 , A61K48/00 , C12N15/102 , C12N15/8213 , C12N2310/315 , C12N2310/321 , C12N2310/3231 , C12N2310/346 , Y02A50/473 , C12N2310/3521
摘要： Presented are methods and compositions for targeted chromosomal genomic alterations using modified single-stranded oligonucleotides. The oligonucleotides of the invention have at least one modified nuclease-resistant terminal region comprising phosphorothioate linkages, LNA analogs or 2′-O-Me base analogs.
摘要翻译：提出了使用修饰的单链寡核苷酸的靶向染色体基因组改变的方法和组合物。本发明的寡核苷酸具有包含硫代磷酸酯键，LNA类似物或2'-O-Me碱基类似物的至少一个修饰的核酸酶抗性末端区域。

4. 发明授权

US07229767B2 Genomics applications for modified OLIGO nucleotides 失效
标题翻译：用于修饰的OLIGO核苷酸的基因组学应用
公开(公告)号：US07229767B2
公开(公告)日：2007-06-12
申请号：US10672735
申请日：2003-09-26
申请人： Eric B. Kmiec , Howard B. Gamper , Michael C. Rice , Michael G. Usher
发明人： Eric B. Kmiec , Howard B. Gamper , Michael C. Rice , Michael G. Usher
IPC分类号： C07H21/04 , C12P19/34 , C12Q1/68
CPC分类号： C12Q1/6813 , C12N15/101 , C12Q1/6816 , C12Q2525/113 , C12Q2521/507
摘要： Methods for the production and use of stable complexes of duplex nucleic acid molecules and oligonucleotides are presented. These complexes can be used for the detection and purification of a known nucleic acid target as well as the manipulation of a defined nucleic acid target sequence.
摘要翻译：提出了生产和使用双链核酸分子和寡核苷酸的稳定复合物的方法。这些配合物可以用于检测和纯化已知的核酸靶，以及操作所定义的核酸靶序列。

5. 发明授权

US5935830A Targeted mutagenesis in living cells using modified oligonucleotides 失效
公开(公告)号：US5935830A
公开(公告)日：1999-08-10
申请号：US827116
申请日：1997-03-26
申请人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
发明人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
IPC分类号： A61K31/70 , A61K38/00 , A61K48/00 , A61P31/00 , A61P35/00 , C07H21/00 , C12N15/10 , C12N15/113 , C12Q1/68 , C12N15/00 , C07H21/04
CPC分类号： C12N15/113 , C07H21/00 , C12N15/102 , C12Q1/6839 , A61K38/00 , C12N2310/153 , C12N2310/311 , C12N2310/335 , C12N2310/3511
摘要： A method for introducing a site-specific mutation into a target polynucleotide sequence is presented. The method involves the use of an oligonucleotide capable of binding to the target sequence, either by triplex formation (mediated by Hoogsteen, reverse Hoogsteen or equivalent base pairing) or by Watson/Crick base pairing (in the presence of a recombinase enzyme). The oligonucleotide of the invention is modified by the covalent attachment of one or more electrophilic groups. When a modified oligonucleotide is bound to its target sequence, the electrophilic group is able to interact with a nearby nucleotide in the target sequence, causing a modification to the nucleotide that results in a change in nucleotide sequence. Compositions used in the practice of the method are also disclosed.

6. 发明授权

US5659022A Oligonucleotide-cyclopropapyrroloindole conjugates as sequence specific hybridization and crosslinking agents for nucleic acids 失效
标题翻译：寡核苷酸 - 环丙吡咯并吲哚缀合物作为核酸的序列特异性杂交和交联剂
公开(公告)号：US5659022A
公开(公告)日：1997-08-19
申请号：US583435
申请日：1996-01-05
申请人： Igor V. Kutyavin , Eugeny A. Lukhtanov , Howard B. Gamper , Rich B. Meyer, Jr. , Alexander Gall
发明人： Igor V. Kutyavin , Eugeny A. Lukhtanov , Howard B. Gamper , Rich B. Meyer, Jr. , Alexander Gall
IPC分类号： C07H19/044 , C07H21/00 , C07H19/00 , A01N43/04 , C07H21/02 , C07H21/04
CPC分类号： C07H21/00 , C07H19/044
摘要： Covalently linked conjugates of oligonucleotides (ODNs) with a cyclopropapyrroloindole moiety or an analog thereof, selectively and efficiently alkylate and crosslink with nucleic acid sequences that are complementary to the base sequence of the ODN.
摘要翻译：寡核苷酸（ODN）与环丙吡咯并吲哚部分或其类似物的共价连接的缀合物选择性且有效地与与ODN的碱基序列互补的核酸序列烷基化和交联。

7. 发明授权

US06312953B1 Bifunctional Crosslinking oligonucleotides adapted for linking to a target sequence of duplex DNA 失效
标题翻译：双功能交联寡核苷酸适于与双链DNA的靶序列连接
公开(公告)号：US06312953B1
公开(公告)日：2001-11-06
申请号：US08266949
申请日：1994-06-27
申请人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
发明人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
IPC分类号： C12Q168
CPC分类号： C07H21/00
摘要： Chemically modified oligonucleotides (ODNS) are complementary, either in the sense of the classic “four letter code” recognition motif, or in the sense required for triple strand formation based on the more limited “two letter code recognition motif”, to a target sequence of double stranded DNA of an invading cell, organism or pathogen, such as a virus, fungus, parasite, bacterium, malignant cell, or any duplex DNA which is desired to be broken into segments for the purpose of “mapping”. The ODNs have cross-linking agents covalently attached at least to two different sites of the ODN. Alternatively, the cross-linking agent which is attached to one site on the ODN has two cross-linking functionalities, and therefore in effect comprises two cross-linking agents. The cross-linking agent typically includes a linker arm (such as an alkyl, alkoxy, aminoalkyl or amidoalkyl chain) and a reactive group which, after triple strand formation with the target sequence of DNA, is capable of reacting with the target DNA to form a covalent bond therewith. Each cross-linking agent of the novel modified ODNs is capable of forming a covalent bond with the target DNA. As a result of the covalent bond formation between the modified ODN and both strands of the target DNA sequence, replication and expression of the target DNA sequence is inhibited. Alternatively the duplex DNA is selectively cleaved with enzymes or amino acids, at the alkylation sites for “mapping” or other investigative purposes.
摘要翻译：化学修饰的寡核苷酸（ODNS）在经典的“四字母代码”识别基序的意义上是互补的，或者在基于更有限的“双字母代码识别基序”的三链形成所需的意义上与目标序列的入侵细胞，生物体或病原体如病毒，真菌，寄生虫，细菌，恶性细胞或任何双链DNA的双链DNA，其为了“映射”的目的而被分解成片段。 ODN具有共价连接到ODN的至少两个不同位点的交联剂。或者，连接到ODN上的一个位点的交联剂具有两个交联官能团，因此实际上包含两种交联剂。交联剂通常包括连接臂（例如烷基，烷氧基，氨基烷基或酰氨基烷基链）和反应性基团，其在与DNA的靶序列形成三链后能够与靶DNA反应形成与其共价键。新型修饰的ODN的交联剂能与靶DNA形成共价键。由于修饰的ODN与目标DNA序列的两条链之间共价键的形成，靶DNA序列的复制和表达被抑制。或者，双链DNA在烷基化位点被选择性地用酶或氨基酸切割，用于“映射”或其它调查目的。

8. 发明授权

US6136601A Targeted mutagenesis in living cells using modified oligonucleotides 失效
标题翻译：使用修饰的寡核苷酸在活细胞中靶向诱变
公开(公告)号：US6136601A
公开(公告)日：2000-10-24
申请号：US827117
申请日：1997-03-26
申请人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
发明人： Rich B. Meyer, Jr. , Howard B. Gamper , Igor V. Kutyavin , Alexander A. Gall
IPC分类号： C07H21/00 , C12Q1/68 , C07H21/04
CPC分类号： C07H21/00 , C12Q1/6839
摘要： A method for introducing a site-specific mutation into a target polynucleotide sequence is presented. The method involves the use of an oligonucleotide capable of binding to the target sequence, either by triplex formation (mediated by Hoogsteen, reverse Hoogsteen or equivalent base pairing) or by Watson/Crick base pairing (in the presence of a recombinase enzyme). The oligonucleotide of the invention is modified by the covalent attachment of one or more electrophilic groups. When a modified oligonucleotide is bound to its target sequence, the electrophilic group is able to interact with a nearby nucleotide in the target sequence, causing a modification to the nucleotide that results in a change in nucleotide sequence. Compositions used in the practice of the method are also disclosed.
摘要翻译：提出了将位点特异性突变引入靶多核苷酸序列的方法。该方法包括通过三重形成（由Hoogsteen，逆Hoogsteen或等效碱基配对介导）或通过Watson / Crick碱基配对（在重组酶存在下）使用能够结合靶序列的寡核苷酸。本发明的寡核苷酸通过一个或多个亲电子基团的共价连接被修饰。当修饰的寡核苷酸与其靶序列结合时，亲电子基团能够与靶序列中的附近核苷酸相互作用，导致导致核苷酸序列变化的核苷酸修饰。还公开了在该方法的实践中使用的组合物。

9. 发明授权

US09969765B2 Nuclions and ribocapsids 有权
公开(公告)号：US09969765B2
公开(公告)日：2018-05-15
申请号：US13883918
申请日：2011-11-07
申请人： Nigel L. Webb , Howard B. Gamper
发明人： Nigel L. Webb , Howard B. Gamper
IPC分类号： C07H21/02 , C12Q1/68 , C12P19/34
CPC分类号： C07H21/02 , C12Q1/6883 , C12Q1/6886 , C12Q2600/156
摘要： The invention relates to an isolated nuclion having (i) a core nucleic acid, and (ii) one or more ribocapsids each including a polymer of two or more ribocapsid subunits, wherein said ribocapsid subunits include nucleic acid. The invention also relates to a method for manufacturing an isolated nuclion.

10. 发明申请

US20130267695A1 Nuclions and Ribocapsids 有权
标题翻译：骨髓和核糖体
公开(公告)号：US20130267695A1
公开(公告)日：2013-10-10
申请号：US13883918
申请日：2011-11-07
申请人： Nigel L. Webb , Howard B. Gamper
发明人： Nigel L. Webb , Howard B. Gamper
IPC分类号： C07H21/02
CPC分类号： C07H21/02 , C12Q1/6883 , C12Q1/6886 , C12Q2600/156
摘要： The invention relates to an isolated nuclion having (i) a core nucleic acid, and (ii) one or more ribocapsids each including a polymer of two or more ribocapsid subunits, wherein said ribocapsid subunits include nucleic acid. The invention also relates to a method for manufacturing an isolated nuclion.
摘要翻译：本发明涉及具有（i）核心核酸的分离的核酸，和（ii）一个或多个核糖体，每个核糖包括两个或多个核糖核酸亚单位的聚合物，其中所述核糖核酸亚单位包括核酸。本发明还涉及一种用于制造分离的核酸的方法。

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