公开(公告)号：US07700758B2

公开(公告)日：2010-04-20

申请号：US10622240

申请日：2003-07-18

Applicant: George Tzertzinis ,  George Feehery ,  Christopher Noren ,  Corinna Tuckey ,  Larry McReynolds ,  Yinhua Zhang

Inventor： George Tzertzinis ,  George Feehery ,  Christopher Noren ,  Corinna Tuckey ,  Larry McReynolds ,  Yinhua Zhang

IPC: C07N21/04 ,  C12Q1/68 ,  C12N5/00 ,  A61K31/70

CPC classification number: C12N15/111 ,  C12N2310/14 ,  C12N2330/30 ,  C12N2330/31 ,  C12Y301/26003

Abstract: A method for obtaining a mixture of heterogenous short double-stranded RNA molecules suitable for use in gene silencing (hsiRNA) by subjecting large double-stranded RNA to enzymatic cleavage under specified conditions. The resulting mixture consistently includes enhanced representation of fragments having a size of 21-22 nucleotides absent any fractionation step. The fragments contain sequences that collectively span the entire length of the large double-stranded RNA from which they are derived. Double-stranded RNA with sequences that individually represent segments of a target mRNA may be analyzed using the methods described herein to identify the most active subset of hsiRNA fragments or individual siRNA fragments for achieving gene silencing for any gene or transcribed sequences. A method is additionally provided for preparing and cloning DNA encoding selected siRNA, hsiRNA mixtures or hairpin sequences to provide a continuous supply of a gene silencing reagent derived from any long double-stranded RNA.

Abstract translation: 一种通过在特定条件下使大的双链RNA进行酶裂解获得适用于基因沉默（hsiRNA）的异源短双链RNA分子的混合物的方法。 所得混合物始终包括具有21-22个核苷酸大小的片段的增强表示，不存在任何分级步骤。 片段含有共同跨越它们衍生的大双链RNA的整个长度的序列。 可以使用本文所述的方法分析具有单独表示靶mRNA的片段的序列的双链RNA，以鉴定用于实现任何基因或转录序列的基因沉默的hsiRNA片段或单个siRNA片段的最活跃子集。 另外提供了用于制备和克隆编码所选siRNA，hsiRNA混合物或发夹序列的DNA以提供从任何长双链RNA衍生的基因沉默试剂的连续供应的方法。

公开(公告)号：US20090133136A1

公开(公告)日：2009-05-21

申请号：US11814158

申请日：2006-01-18

Applicant: Demin Zhou ,  Jon E. Chatterton ,  Ning Ke ,  Jing Zhang ,  Joshua Bliesath ,  Henry Li ,  Flossie Wong-Staal

Inventor： Demin Zhou ,  Jon E. Chatterton ,  Ning Ke ,  Jing Zhang ,  Joshua Bliesath ,  Henry Li ,  Flossie Wong-Staal

IPC: A01K67/027 ,  C12N15/82 ,  C12N15/85 ,  A61K48/00 ,  C40B40/06 ,  C12N5/10

CPC classification number: C12N15/86 ,  A01K2267/0331 ,  C12N15/111 ,  C12N15/113 ,  C12N2310/111 ,  C12N2310/14 ,  C12N2320/10 ,  C12N2320/30 ,  C12N2320/50 ,  C12N2330/31 ,  C12N2740/15043 ,  C12N2830/006 ,  C12N2840/203

Abstract: An inducible siRNA expression polynucleotide and methods for its use are provided. The expression polynucleotide comprises a bicistronic expression cassette that encodes a repressor and a detectable marker, wherein the repressor controls expression of siRNA expression in the absence of an inducer.

Abstract translation: 提供可诱导的siRNA表达多核苷酸及其使用方法。 表达多核苷酸包含编码阻抑物和可检测标记的双顺反子表达盒，其中所述阻遏物在不存在诱导物的情况下控制siRNA表达的表达。

公开(公告)号：US20090105174A1

公开(公告)日：2009-04-23

申请号：US12105562

申请日：2008-04-18

Applicant: Sumedha D. Jayasena

Inventor： Sumedha D. Jayasena

IPC: A61K31/7105 ,  C12N5/06 ,  C07H21/02 ,  C12N5/08

CPC classification number: C12N15/113 ,  C12N2310/11 ,  C12N2310/315 ,  C12N2310/321 ,  C12N2310/3231 ,  C12N2310/3515 ,  C12N2320/11 ,  C12N2330/31 ,  C12N2310/3521

Abstract: Methods and compositions relating to nucleic acids targeting certain miRNA molecules are disclosed. The nucleic acids are useful in methods of increasing nuclear concentration of FKHR protein, decreasing cell viability, and treating cancer.

Abstract translation: 公开了涉及靶向某些miRNA分子的核酸的方法和组合物。 核酸可用于增加FKHR蛋白的核浓度，降低细胞活力和治疗癌症的方法。

公开(公告)号：US20090099043A1

公开(公告)日：2009-04-16

申请号：US12313554

申请日：2008-11-21

Applicant: York Yuan Yuan Zhu

Inventor： York Yuan Yuan Zhu

IPC: C40B50/06 ,  C07H21/00 ,  C12N15/63

CPC classification number: C12N15/113 ,  C12N15/111 ,  C12N2310/14 ,  C12N2330/31 ,  C40B40/08 ,  C40B50/06

Abstract: The present invention provides a PCR based high-throughput method for preparing full-sites siRNA polynucleotide pool, comprising: DNase I random digestion; Loop-1 phosphate linker ligation; single PCR amplification; a type III restriction/modification enzyme digestion; blunt ending; Loop-2 phosphate linker ligation; double primer PCR; FokI digestion and cloning into an siRNA expression vector. The present invention enables the use of a type III restriction/modification enzyme linkers mediated PCR method for high-throughput preparing an siRNA polynucleotide pool, in which the functional length of siRNAs can be controllably distributed from 19-23 bp, thus completely mimic the natural siRNA length diversity, specially suitable for RNAi therapeutic targets screening. The present invention overcomes the bottlenecks and drawbacks of conventional siRNA polynucleotide pool construction technologies.

Abstract translation: 本发明提供了一种用于制备全位点siRNA多核苷酸库的基于PCR的高通量方法，其包括：DNase I随机消化; 环磷酸酯接头连接; 单PCR扩增; III型限制/修饰酶消化; 钝的结局 环2磷酸酯接头连接; 双引物PCR; FokI消化并克隆入siRNA表达载体。 本发明能够使用III型限制性/修饰酶接头介导的PCR方法，用于高通量制备siRNA多核苷酸库，其中siRNA的功能长度可以从19-23bp可控地分布，从而完全模拟天然 siRNA长度多样性，特别适用于RNAi治疗靶点筛选。 本发明克服了常规siRNA多核苷酸库构建技术的瓶颈和弊端。

公开(公告)号：US20080213861A1

公开(公告)日：2008-09-04

申请号：US11894676

申请日：2007-08-20

Applicant: Gregory J. Hannon ,  Patrick J. Paddison ,  Despina C. Siolas

Inventor： Gregory J. Hannon ,  Patrick J. Paddison ,  Despina C. Siolas

IPC: C12N9/18

CPC classification number: C12N15/113 ,  A01K2217/05 ,  A61K38/00 ,  C12N15/1034 ,  C12N15/1079 ,  C12N15/111 ,  C12N2310/111 ,  C12N2310/14 ,  C12N2310/53 ,  C12N2320/12 ,  C12N2330/30 ,  C12N2330/31 ,  C12Y301/26003

Abstract: The present invention provides methods for attenuating gene expression in a cell, especially in a mammalian cell, using gene-targeted double stranded RNA (dsRNA), such as a hairpin RNA. The dsRNA contains a nucleotide sequence that hybridizes under physiologic conditions of the cell to the nucleotide sequence of at least a portion of the gene to be inhibited (the “target” gene).

Abstract translation: 本发明提供使用基因靶向双链RNA（dsRNA）如发夹RNA来减弱细胞，特别是哺乳动物细胞中的基因表达的方法。 dsRNA含有在细胞的生理条件下与要抑制的基因的至少一部分（“靶”基因）的核苷酸序列杂交的核苷酸序列。

公开(公告)号：US20070218495A1

公开(公告)日：2007-09-20

申请号：US11724346

申请日：2007-03-15

Applicant: Amanda Birmingham ,  Emily Anderson ,  Angela Reynolds ,  Devin Leake ,  Scott Baskerville ,  Yuriy Fedorov ,  Jon Karpilow ,  William Marshall ,  Anastasia Khvorova

Inventor： Amanda Birmingham ,  Emily Anderson ,  Angela Reynolds ,  Devin Leake ,  Scott Baskerville ,  Yuriy Fedorov ,  Jon Karpilow ,  William Marshall ,  Anastasia Khvorova

IPC: C40B40/08 ,  C07H21/02

CPC classification number: C12N15/111 ,  C12N2310/14 ,  C12N2320/50 ,  C12N2330/31

Abstract: The present invention provides methods, libraries and computer program products for selecting siRNA that reduce off-target effects and methods for gene silencing using these siRNAs. By comparing nucleotide sequences at positions 2-7 or 2-8 of the sense and/or antisense regions of candidate siRNAs to the 3′ UTR region of mRNAs, one can select siRNAs that have reduced off-target effects.

Abstract translation: 本发明提供用于选择减少脱靶效应的siRNA的方法，文库和计算机程序产品以及使用这些siRNA的基因沉默方法。 通过将候选siRNAs的有义和/或反义区的2-7位或2-8位的核苷酸序列与mRNA的3'UTR区进行比较，可以选择具有降低的脱靶效应的siRNA。

公开(公告)号：US20070027103A1

公开(公告)日：2007-02-01

申请号：US11542291

申请日：2006-10-03

Applicant: Catherine Pachuk

Inventor： Catherine Pachuk

IPC: A61K48/00 ,  C12Q1/68 ,  C40B40/08

CPC classification number: C12N15/111 ,  C12N15/1137 ,  C12N2310/13 ,  C12N2310/14 ,  C12N2310/53 ,  C12N2320/12 ,  C12N2320/50 ,  C12N2330/31 ,  C12Y301/03001

Abstract: The present invention provides methods of post-transcriptional gene silencing which involve the use of a first dsRNA having substantial sequence identity to a target nucleic acid and a short, second dsRNA which inhibits dsRNA-mediated toxicity. These methods can be used to prevent or treat a disease or infection by silencing a gene associated with the disease or infection. The invention also provides methods for identifying nucleic acid sequences that modulate a detectable phenotype, including the function of a cell, the expression of a gene, or the biological activity of a target polypeptide.

Abstract translation: 本发明提供转录后基因沉默的方法，其涉及使用与靶核酸具有实质序列同一性的第一dsRNA和抑制dsRNA介导的毒性的短的第二dsRNA。 这些方法可用于通过沉默与该疾病或感染相关的基因来预防或治疗疾病或感染。 本发明还提供了用于鉴定调节可检测表型的核酸序列的方法，所述核酸序列包括细胞的功能，基因的表达或靶多肽的生物活性。

公开(公告)号：US20060135456A1

公开(公告)日：2006-06-22

申请号：US10997086

申请日：2004-11-23

Applicant: Gregory Hannon ,  Patrick Paddison ,  Despina Siolas

Inventor： Gregory Hannon ,  Patrick Paddison ,  Despina Siolas

IPC: A61K48/00

CPC classification number: C12N15/113 ,  A01K2217/05 ,  A61K38/00 ,  C12N15/1034 ,  C12N15/1079 ,  C12N15/111 ,  C12N2310/111 ,  C12N2310/14 ,  C12N2310/53 ,  C12N2320/12 ,  C12N2330/30 ,  C12N2330/31 ,  C12Y301/26003

Abstract: The present invention provides methods for attenuating gene expression in a cell, especially in a mammalian cell, using gene-targeted double stranded RNA (dsRNA), such as a hairpin RNA. The dsRNA contains a nucleotide sequence that hybridizes under physiologic conditions of the cell to the nucleotide sequence of at least a portion of the gene to be inhibited (the “target” gene).

Abstract translation: 本发明提供使用基因靶向双链RNA（dsRNA）如发夹RNA来减弱细胞，特别是哺乳动物细胞中的基因表达的方法。 dsRNA含有在细胞的生理条件下与要抑制的基因的至少一部分（“靶”基因）的核苷酸序列杂交的核苷酸序列。

公开(公告)号：US20050260652A1

公开(公告)日：2005-11-24

申请号：US11106238

申请日：2005-04-14

Applicant: Gary Ruvkun ,  Ravi Kamath ,  Harrison Gabel ,  John Kim

Inventor： Gary Ruvkun ,  Ravi Kamath ,  Harrison Gabel ,  John Kim

IPC: A01K67/033 ,  C12N15/11 ,  C12Q1/68

CPC classification number: C12N15/111 ,  A01K2217/05 ,  A01K2217/075 ,  A01K2227/105 ,  C12N2310/14 ,  C12N2320/12 ,  C12N2320/50 ,  C12N2330/31

Abstract: The invention features compositions useful in modulating RNA interference in a wide variety of cell types for the treatment of virtually any disease or disorder related to the overexpression of a gene or genes, as well as methods of identifying and using such compositions.

Abstract translation: 本发明的特征在于可用于调节多种细胞类型中的RNA干扰的组合物，用于治疗与基因或基因的过表达相关的任何疾病或病症，以及鉴定和使用这些组合物的方法。

公开(公告)号：US20050222067A1

公开(公告)日：2005-10-06

申请号：US10968821

申请日：2004-10-19

Applicant: Sebastien Pfeffer ,  Thomas Tuschl

Inventor： Sebastien Pfeffer ,  Thomas Tuschl

IPC: A61K48/00 ,  C07H21/04 ,  C07K14/045 ,  C12N15/11 ,  C12N15/113 ,  C12N15/86

CPC classification number: C12N15/1131 ,  C07H21/04 ,  C07K14/005 ,  C12N15/111 ,  C12N2310/111 ,  C12N2310/14 ,  C12N2310/53 ,  C12N2320/11 ,  C12N2330/10 ,  C12N2330/31 ,  C12N2710/16022 ,  C12N2710/16222 ,  C12N2710/16422

Abstract: The invention relates to isolated nucleic acid molecules comprising the sequence of a human cytomegalovirus microRNA. In another embodiment, the invention relates to single stranded DNA virus microRNA molecules comprising the sequence of a human cytomegalovirus microRNA. The invention also relates to the anti-DNA virus microRNA molecules.

Abstract translation: 本发明涉及包含人巨细胞病毒微RNA序列的分离的核酸分子。 在另一个实施方案中，本发明涉及包含人巨细胞病毒微RNA序列的单链DNA病毒微小RNA分子。 本发明还涉及抗DNA病毒微小RNA分子。