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1. 发明授权

US11053536B2 Tag-sequence-attached two-dimensional CDNA library device, and gene expression analysis method and gene expression analysis apparatus each utilizing same 有权
公开(公告)号：US11053536B2
公开(公告)日：2021-07-06
申请号：US14418135
申请日：2012-07-30
申请人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi , Maiko Tanabe
发明人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi , Maiko Tanabe
IPC分类号： C12Q1/6837 , C12N15/10
摘要： The present invention relates to a method, a device, and an apparatus for analyzing the expression of a gene in single cells. Specifically, the present invention relates to: a device for gene expression analysis, characterized by including a support, in which a nucleic acid probe having a test nucleic acid capture sequence and a known sequence, and further containing a cell recognition tag sequence which differs depending on the difference in position on the surface of the support or in the vicinity of the surface thereof, and a common primer sequence having a known sequence is two-dimensionally distributed and immobilized on the surface of the support or in the vicinity of the surface thereof; and a method and an apparatus using the device for gene expression analysis.

2. 发明申请

US20150167063A1 Tag-Sequence-Attached Two-Dimensional CDNA Library Device, and Gene Expression Analysis Method and Gene Expression Analysis Apparatus Each Utilizing Same 审中-公开
标题翻译：标签序列连接的二维CDNA文库装置及基因表达分析方法及基因表达分析装置
公开(公告)号：US20150167063A1
公开(公告)日：2015-06-18
申请号：US14418135
申请日：2012-07-30
申请人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi , Maiko Tanabe
发明人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi , Maiko Tanabe
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6837 , C12N15/1096 , C12Q2525/155 , C12Q2565/537 , C12Q2565/601
摘要： The present invention relates to a method, a device, and an apparatus for analyzing the expression of a gene in single cells. Specifically, the present invention relates to: a device for gene expression analysis, characterized by including a support, in which a nucleic acid probe having a test nucleic acid capture sequence and a known sequence, and further containing a cell recognition tag sequence which differs depending on the difference in position on the surface of the support or in the vicinity of the surface thereof, and a common primer sequence having a known sequence is two-dimensionally distributed and immobilized on the surface of the support or in the vicinity of the surface thereof; and a method and an apparatus using the device for gene expression analysis.
摘要翻译：本发明涉及用于分析单细胞中基因表达的方法，装置和装置。具体地说，本发明涉及：用于基因表达分析的装置，其特征在于，包括载体，其中具有测试核酸捕获序列和已知序列的核酸探针，并且还包含不同的细胞识别标签序列关于载体表面或其表面附近的位置差异，具有已知序列的常见引物序列被二维分布和固定在载体的表面上或其表面附近 ; 以及使用该基因表达分析装置的方法和装置。

3. 发明申请

US20120245053A1 GENE EXPRESSION ANALYSIS METHOD USING TWO DIMENSIONAL cDNA LIBRARY 审中-公开
标题翻译：使用两维cDNA文库的基因表达分析方法
公开(公告)号：US20120245053A1
公开(公告)日：2012-09-27
申请号：US13513605
申请日：2010-11-29
申请人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi
发明人： Masataka Shirai , Hideki Kambara , Kiyomi Taniguchi
IPC分类号： C40B30/04
CPC分类号： C12N15/1093 , C12Q1/6809 , C12Q1/6837 , C40B40/08 , C40B50/06 , G01N33/54306 , G01N2458/10 , C12Q2531/125 , C12Q2533/107 , C12Q2565/537
摘要： The present invention provides a method and/or means for collecting and analyzing an individual cell in a tissue, and at the same time, quantitatively monitoring the expression levels of various genes while keeping two-dimensional information in the tissue. Specifically, the present invention provides a method comprising preparing a cDNA library from mRNA while keeping two-dimensional cellular distribution information and obtaining the gene expression levels at any site or all sites at a level of single cell. More specifically, the present invention provides a method comprising preparing a cDNA library in a sheet-form from mRNA while keeping two-dimensional cellular distribution information and repeatedly using the cDNA library in the detection of the gene expression, thereby allowing measurement of the expression distribution for a number of genes at a high accuracy.
摘要翻译：本发明提供了一种用于收集和分析组织中的单个细胞的方法和/或方法，并且同时定量监测各种基因的表达水平，同时保持组织中的二维信息。具体地说，本发明提供一种方法，其包括从mRNA制备cDNA文库，同时保持二维细胞分布信息，并在单细胞水平的任何位点或所有位点获得基因表达水平。更具体地，本发明提供了一种方法，其包括从mRNA制备片段的cDNA文库，同时保持二维细胞分布信息，并在检测基因表达中重复使用cDNA文库，从而允许测量表达分布对于许多基因在高精度。

4. 发明授权

US08389246B2 Method for nucleic acid quantitation 失效
公开(公告)号：US08389246B2
公开(公告)日：2013-03-05
申请号：US12292800
申请日：2008-11-26
申请人： Kiyomi Taniguchi , Hideki Kambara
发明人： Kiyomi Taniguchi , Hideki Kambara
IPC分类号： C12P19/34
CPC分类号： C12Q1/6851 , C12Q2545/101 , C12Q2525/161
摘要： It is intended to provide a novel convenient approach for DNA quantitative analysis that overcomes the disadvantages of conventional formulations. A standard DNA sample is prepared by introducing a single-base substitution into target DNA, and a predetermined amount thereof is mixed with a target DNA sample. The target and standard DNAs are amplified using the same primers designed to amplify a region comprising the single-base substitution site. To a hybridization product of a probe capable of binding to a site immediately before the single-base substitution site, ddATP, ddGTP, ddCTP, and ddTTP are sequentially added one by one to perform a complementary strand synthesis reaction. Luciferase reaction-induced luminescence derived from the formed pyrophosphoric acid is detected. The target DNA is quantitated from the amount of the detected luminescence and the amount of the added standard DNA sample.

5. 发明申请

US20090142767A1 Method for nucleic acid quantitation 失效
标题翻译：核酸定量方法
公开(公告)号：US20090142767A1
公开(公告)日：2009-06-04
申请号：US12292800
申请日：2008-11-26
申请人： Kiyomi Taniguchi , Hideki Kambara
发明人： Kiyomi Taniguchi , Hideki Kambara
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6851 , C12Q2545/101 , C12Q2525/161
摘要： It is intended to provide a novel convenient approach for DNA quantitative analysis that overcomes the disadvantages of conventional formulations. A standard DNA sample is prepared by introducing a single-base substitution into target DNA, and a predetermined amount thereof is mixed with a target DNA sample. The target and standard DNAs are amplified using the same primers designed to amplify a region comprising the single-base substitution site. To a hybridization product of a probe capable of binding to a site immediately before the single-base substitution site, ddATP, ddGTP, ddCTP, and ddTTP are sequentially added one by one to perform a complementary strand synthesis reaction. Luciferase reaction-induced luminescence derived from the formed pyrophosphoric acid is detected. The target DNA is quantitated from the amount of the detected luminescence and the amount of the added standard DNA sample.
摘要翻译：旨在为DNA定量分析提供一种新颖的方便方法，克服了常规制剂的缺点。通过将单碱基置换引入目标DNA中制备标准DNA样品，并将其预定量与目标DNA样品混合。使用设计用于扩增包含单碱基取代位点的区域的相同引物扩增靶标和标准DNA。对于能够在单碱基置换位点之前结合位点的探针的杂交产物，逐个依次加入ddATP，ddGTP，ddCTP和ddTTP以进行互补链合成反应。检测到由形成的焦磷酸衍生的荧光素酶反应诱导的发光。从所检测的发光量和添加的标准DNA样品的量定量靶DNA。

6. 发明申请

US20070281313A1 Methods for quantitative cDNA analysis in single-cell 有权
标题翻译：单细胞定量cDNA分析方法
公开(公告)号：US20070281313A1
公开(公告)日：2007-12-06
申请号：US11783575
申请日：2007-04-10
申请人： Kiyomi Taniguchi , Hideki Kambara , Tomoharu Kajiyama
发明人： Kiyomi Taniguchi , Hideki Kambara , Tomoharu Kajiyama
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6811 , C12N15/1096 , C12Q1/6809 , C12Q1/6834 , C12Q2565/537 , C12Q2525/173 , C12Q2521/301 , C12Q2561/113
摘要： It is an object to provide a method of suitably analyzing the amount of gene expression of a single-cell.A method of detecting a nucleic acid comprisinga step of sampling a single-cell from a sample containing at least a single-cell,a cell lysis step of lysing cell membrane of the sampled single-cell and extracting nucleic acids from the cell,a DNase treatment step of degrading DNA of the extracted nucleic acids with DNase,a step of hybridizing mRNA of the total RNA contained in the single-cell with oligo (dT) fixed onto a carrier,a step of performing reverse transcription of the mRNA hybridized with the oligo (dT) to fix cDNA derived from the single-cell onto the carrier, thereby preparing a single-cell derived cDNA library fixed onto a carrier, anda step of amplifying cDNA fixed onto the carrier and simultaneously detecting an amplification amount of the cDNA.
摘要翻译：本发明的目的是提供适当分析单细胞的基因表达量的方法。一种检测核酸的方法，包括从含有至少单细胞的样品中取样单细胞的步骤，裂解采样的单细胞的细胞膜并从细胞中提取核酸的细胞裂解步骤用DNase降解提取的核酸的DNA的DNA酶处理步骤，将单细胞中含有的总RNA的mRNA与固定在载体上的寡聚（dT）杂交的步骤，将与mRNA杂交的mRNA进行逆转录的步骤将来自单细胞的cDNA固定在载体上的寡核苷酸（dT），从而制备固定在载体上的单细胞衍生的cDNA文库，以及扩增固定在载体上的cDNA并同时检测扩增量的步骤 cDNA。

7. 发明授权

US08802367B2 Methods for quantitative cDNA analysis in single-cell 有权
标题翻译：单细胞定量cDNA分析方法
公开(公告)号：US08802367B2
公开(公告)日：2014-08-12
申请号：US11783575
申请日：2007-04-10
申请人： Kiyomi Taniguchi , Hideki Kambara , Tomoharu Kajiyama
发明人： Kiyomi Taniguchi , Hideki Kambara , Tomoharu Kajiyama
IPC分类号： C12Q1/68 , C12P19/34 , C07H21/02 , C07H21/04
CPC分类号： C12Q1/6811 , C12N15/1096 , C12Q1/6809 , C12Q1/6834 , C12Q2565/537 , C12Q2525/173 , C12Q2521/301 , C12Q2561/113
摘要： It is an object to provide a method of suitably analyzing the amount of gene expression of a single-cell.A method of detecting a nucleic acid comprising a step of sampling a single-cell from a sample containing at least a single-cell, a cell lysis step of lysing cell membrane of the sampled single-cell and extracting nucleic acids from the cell, a DNase treatment step of degrading DNA of the extracted nucleic acids with DNase, a step of hybridizing mRNA of the total RNA contained in the single-cell with oligo (dT) fixed onto a carrier, a step of performing reverse transcription of the mRNA hybridized with the oligo (dT) to fix cDNA derived from the single-cell onto the carrier, thereby preparing a single-cell derived cDNA library fixed onto a carrier, and a step of amplifying cDNA fixed onto the carrier and simultaneously detecting an amplification amount of the cDNA.
摘要翻译：本发明的目的是提供适当分析单细胞的基因表达量的方法。一种检测核酸的方法，包括从含有至少单细胞的样品中取样单细胞的步骤，裂解采样的单细胞的细胞膜并从细胞中提取核酸的细胞裂解步骤用DNase降解提取的核酸的DNA的DNA酶处理步骤，将单细胞中含有的总RNA的mRNA与固定在载体上的寡聚（dT）杂交的步骤，将与mRNA杂交的mRNA进行逆转录的步骤将来自单细胞的cDNA固定在载体上的寡核苷酸（dT），从而制备固定在载体上的单细胞衍生的cDNA文库，以及扩增固定在载体上的cDNA并同时检测扩增量的步骤 cDNA。

8. 发明申请

US20060223087A1 Method for nucleic acid analysis 审中-公开
标题翻译：核酸分析方法
公开(公告)号：US20060223087A1
公开(公告)日：2006-10-05
申请号：US11342861
申请日：2006-01-31
申请人： Kiyomi Taniguchi , Keiichi Nagai , Hiroko Matsunaga
发明人： Kiyomi Taniguchi , Keiichi Nagai , Hiroko Matsunaga
IPC分类号： C12Q1/68
CPC分类号： C12Q1/683 , C12Q2537/113 , C12Q2523/125 , C12Q2521/307
摘要： A simple and highly accurate method for detecting the presence or absence of gene mutation and methylated cytosine in CpG dinucleotide that are contained in a target sequence derived from an analysis sample is provided. Features of the method for nucleic acid analysis include cleaving one or more noncomplementary sites in a double-stranded nucleic acid sample by a single strand-specific endonuclease, hybridizing at least one of the nucleic acid fragments obtained to a probe containing a nucleotide sequence that is partially or totally identical to either one strand of the double-stranded nucleic acid sample, allowing an extension reaction to proceed from the nucleic acid fragment hybridized to the probe, and optically detecting pyrophosphate generated by the extension reaction, thereby judging the presence or absence of at least a noncomplementary site in the double-stranded nucleic acid sample.
摘要翻译：提供了一种用于检测来自分析样品的靶序列中包含的CpG二核苷酸中基因突变和甲基化胞嘧啶的存在或不存在的简单且高度准确的方法。用于核酸分析的方法的特征包括通过单链特异性内切核酸酶切割双链核酸样品中的一个或多个非互补位点，将获得的核酸片段中的至少一个与包含核苷酸序列的探针杂交部分或全部与双链核酸样品的一条链相同，允许从与探针杂交的核酸片段进行延伸反应，并光学检测由延伸反应产生的焦磷酸盐，从而判断是否存在至少在双链核酸样品中的非互补位点。

9. 发明申请

US20050287549A1 Method of genetic testing 审中-公开
标题翻译：基因检测方法
公开(公告)号：US20050287549A1
公开(公告)日：2005-12-29
申请号：US11041456
申请日：2005-01-25
申请人： Keiichi Nagai , Kazunori Okano , Hideyuki Noda , Hiroko Matsunaga , Kiyomi Taniguchi , Yoshiaki Yazawa , Tomoharu Kajiyama
发明人： Keiichi Nagai , Kazunori Okano , Hideyuki Noda , Hiroko Matsunaga , Kiyomi Taniguchi , Yoshiaki Yazawa , Tomoharu Kajiyama
IPC分类号： C12Q1/68
CPC分类号： C12Q1/6827 , C12Q1/6837 , C12Q2565/537 , C12Q2565/301 , C12Q2525/155
摘要： This invention provides a method of genetic testing that enables testing of a plurality of variation sites (SNPs) in a cost-effective and simple manner, allowing realization of genetic diagnosis in clinical settings. The SNP type of the nucleic acid sample is evaluated by: allowing a nucleic acid sample having an anchor sequence at its 5′ end to hybridize to a support having, immobilized on its surface, a probe containing a sequence that is complementary to the target sequence (the SNP region); extending a complementary strand from the probe utilizing the nucleic acid sample as a template; dissociating and removing the nucleic acid sample from the extended probe; extending a complementary strand using the extended probe as a template and a primer having a sequence identical to the anchor sequence; and detecting pyrophosphoric acid generated via the primer extension, based on bioluminescence.
摘要翻译：本发明提供了一种能够以成本有效和简单的方式测试多个变异位点（SNP）的遗传测试方法，从而允许在临床环境中实现遗传诊断。核酸样品的SNP类型通过以下方式评估：使具有其5'端的锚定序列的核酸样品与固定在其表面上的载体杂交，所述载体固定有包含与靶序列互补的序列的探针（SNP地区）; 使用核酸样品作为模板从探针延伸互补链; 从扩展的探针中分离并除去核酸样品; 使用扩展探针作为模板和具有与锚定序列相同的序列的引物延伸互补链; 并基于生物发光检测通过引物延伸产生的焦磷酸。

10. 发明授权

US06734557B2 Semiconductor device 失效
标题翻译：半导体器件
公开(公告)号：US06734557B2
公开(公告)日：2004-05-11
申请号：US10369760
申请日：2003-02-21
申请人： Kiyomi Taniguchi , Hisashige Nishida
发明人： Kiyomi Taniguchi , Hisashige Nishida
IPC分类号： H01L2348
CPC分类号： H05K3/3452 , H01L21/4853 , H01L23/49838 , H01L24/48 , H01L2224/0401 , H01L2224/05599 , H01L2224/32225 , H01L2224/48091 , H01L2224/48227 , H01L2224/48464 , H01L2224/73265 , H01L2224/85399 , H01L2924/00014 , H01L2924/01004 , H01L2924/01005 , H01L2924/01006 , H01L2924/01028 , H01L2924/01029 , H01L2924/01078 , H01L2924/01079 , H01L2924/014 , H01L2924/15311 , H01L2924/181 , H05K1/113 , H05K2201/09527 , H05K2201/0989 , H01L2924/00 , H01L2224/45099 , H01L2924/00012 , H01L2224/45015 , H01L2924/207
摘要： A semiconductor device comprising: a substrate; first and second interconnection patterns respectively provided on upper and lower surfaces of the substrate; a through-hole electrode extending through the substrate for electrically connecting the first and second interconnection patterns; a semiconductor chip provided on the upper surface of the substrate and electrically connected to the first interconnection pattern; and a resist film covering the second interconnection pattern; the second interconnection pattern comprising a generally round land and a lead interconnection portion extending from the land, the resist film having an opening formed therein for exposing the entire land, the opening having a curved edge surrounding a peripheral edge of the land and a linear edge linearly extending along a boundary between the land and the lead interconnection portion, the exposed land having a solder ball as an external terminal thereon.
摘要翻译：一种半导体器件，包括：衬底; 分别设置在基板的上表面和下表面上的第一和第二互连图案; 延伸穿过所述基板的用于电连接所述第一和第二互连图案的通孔电极; 设置在所述基板的上表面并电连接到所述第一互连图案的半导体芯片; 以及覆盖所述第二互连图案的抗蚀剂膜; 所述第二互连图案包括大致圆形的平台和从所述焊盘延伸的引线互连部分，所述抗蚀剂膜具有形成在其中的开口，用于暴露所述整个焊盘，所述开口具有围绕所述焊盘边缘的弯曲边缘，沿着焊盘和引线互连部分之间的边界线性延伸，暴露的焊盘在其上具有焊球作为外部端子。

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