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1. 发明申请

US20060257997A1 High throughput synthesis system and synthesizer for automatically performing the system 审中-公开
标题翻译：高吞吐量合成系统和合成器，用于自动执行系统
公开(公告)号：US20060257997A1
公开(公告)日：2006-11-16
申请号：US10554434
申请日：2004-04-23
申请人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara , Riyo Morishita , Mihoro Saeki , Tomohisa Sato , Aya Kitamoto
发明人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara , Riyo Morishita , Mihoro Saeki , Tomohisa Sato , Aya Kitamoto
IPC分类号： C12M3/00
CPC分类号： C12P21/02 , C12P19/34
摘要： To develop system technology for a high throughput in vitro synthesis reaction method for biopolymers such as proteins, RNA, and the like. In detail, a cell-free synthesis system wherein a template substance is used as a raw material, and an automatic synthesizer using the system, includes the following systems, control systems therefor, or combinations thereof; 1) to make a template substance, a substrate, and a reaction solution into contact so as to lead to a synthetic reaction, 2) to put the reaction system outside of the synthetic reaction system to perform dilution treatment of the solution approximately around the decrease in the synthesis velocity, approximately around the stop of the synthetic reaction, or in the middle thereof, 3) to perform concentration treatment subsequent to the dilution treatment, and 4) to put the reaction system back into the synthetic reaction system, or 1) to make a template substance, a substrate, and a reaction solution into contact so as to lead to a synthetic reaction, 2) to put the reaction system outside of the synthetic reaction system to perform concentration treatment of the solution approximately around the decrease in the synthesis velocity, approximately around the stop of the synthetic reaction, or in the middle thereof, 3) to perform dilution treatment subsequent to the concentration treatment, and 4) to put the diluted reaction system back into the synthetic reaction system.
摘要翻译：开发用于生物聚合物如蛋白质，RNA等的高通量体外合成反应方法的系统技术。具体而言，其中使用模板物质作为原料的无细胞合成系统和使用该系统的自动合成器包括以下系统，其控制系统或其组合; 1）使模板物质，底物和反应溶液接触以导致合成反应，2）将反应体系置于合成反应体系外部，以进行大约减少的溶液的稀释处理在合成速度约合成反应停止或其中间，3）进行稀释处理后的浓缩处理，4）使反应体系回到合成反应体系中，或1）使模板物质，底物和反应溶液接触以导致合成反应，2）将反应体系置于合成反应体系外部，以在约合成速度大约在合成反应停止时，或在其中间，3）进行浓缩处理后的稀释处理，和4）将稀释的反应体系放回到合成反应体系中。

2. 发明授权

US07981617B2 Transcription template for cell-free protein synthesis and method using the same 有权
标题翻译：无细胞蛋白质合成的转录模板及其使用方法
公开(公告)号：US07981617B2
公开(公告)日：2011-07-19
申请号：US10363372
申请日：2001-08-28
申请人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara
发明人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara
IPC分类号： C12Q1/68
CPC分类号： C12P21/02 , C12Q1/686
摘要： Methods to construct a transcription template for cell-free protein synthesis that has a high translation template activity, using a 3′-side primer and a 5′-side primer for PCR are provided. The 5′-side primer for PCR has a sequence complementary to a base sequence containing at least a part of a promoter functional site from the 5′-end of a promoter and has a base sequence that does not contain a base sequence complementary to at least a part of a RNA polymerase-recognizing site of the 3′-end of the promoter. The other primer has a base sequence complementary to at least a part of the RNA polymerase-recognizing site of the 3′-end of the promoter and has a sequence that does not contain a base sequence complementary to at least a part of a promoter functional site from the 5′-end of the promoter.
摘要翻译：提供了使用3'侧引物和5'侧引物进行PCR构建具有高翻译模板活性的无细胞蛋白质合成转录模板的方法。用于PCR的5'侧引物具有与包含来自启动子的5'末端的至少一部分启动子功能位点的碱基序列互补的序列，并且其碱基序列不含有与启动子的3'末端的RNA聚合酶识别位点的至少一部分。另一个引物具有与启动子3'末端的RNA聚合酶识别位点的至少一部分互补的碱基序列，并且具有不包含与启动子功能的至少一部分互补的碱基序列的序列来自启动子的5'-末端的位点。

3. 发明授权

US06869774B2 Methods of synthesizing cell-free protein 有权
标题翻译：合成无细胞蛋白的方法
公开(公告)号：US06869774B2
公开(公告)日：2005-03-22
申请号：US10344803
申请日：2001-08-28
申请人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara
发明人： Yaeta Endo , Tatsuya Sawasaki , Tomio Ogasawara
IPC分类号： C12P21/02 , C12P21/06
CPC分类号： C12P21/02
摘要： One embodiment of the present invention is a diffusion continuous batch cell-free protein-synthesis method characterized simultaneously by continuously supplying substrate and energy source molecules in the supply phase to the reaction phase by the free diffusion via interface between both phases and by transferring by-products formed in the reaction phase by enhancing the efficiency of the synthesis reaction by prolonging the reaction lifetime by directly contacting a synthesis reaction mixture (reaction phase) containing a biological extract with a substrate- and energy source-supplying solution (supply phase) without using barrier such as semi-permeable membrane or ultrafiltration membrane in a general cell-free protein-synthesis reaction means. Another embodiment of the present invention is a dilution batch cell-free protein synthesis method characterized by enhancing the efficiency of the protein synthesis by prolonging the reaction lifetime by adding a diluting solution to the reaction mixture after pre-incubating the reaction mixture in a cell-free protein-synthesis reaction means using a wheat-embryo extract. Another embodiment of the present invention is a method characterized by enhancing the efficiency of the synthesis reaction simultaneously by re-supplying substrate and energy sources necessary for the protein synthesis (e.g., amino acids, ATP, GTP, creatine phosphate) to the reaction mixture using a gel filtration column and/or semipermeable membrane and by discontinuously removing by-products formed during the reaction after the synthesis reaction stops in the batch cell-free protein synthesis method.
摘要翻译：本发明的一个实施方案是扩散连续分批无细胞蛋白质合成方法，其特征在于同时通过在两相之间的界面通过自由扩散连续供应反应相中的底物和能量源分子，通过使含有生物提取物的合成反应混合物（反应相）与底物和能量供应溶液（供应阶段）直接接触而延长反应寿命，通过提高合成反应的效率而在反应阶段形成的产物，而不使用一般无细胞蛋白合成反应手段的半透膜或超滤膜等障碍。本发明的另一个实施方案是一种稀释分批无细胞蛋白质合成方法，其特征在于通过在将反应混合物预先培养至细胞培养基中之后，通过向反应混合物中加入稀释溶液来延长反应寿命来提高蛋白质合成的效率，游离蛋白质合成反应意味着使用小麦胚提取物。本发明的另一个实施方案是一种方法，其特征在于通过向反应混合物中再次提供蛋白质合成所需的底物和能量源（例如氨基酸，ATP，GTP，肌酸磷酸），同时提高合成反应的效率，凝胶过滤柱和/或半透膜，并且在间歇无细胞蛋白质合成方法中，在合成反应停止后不连续地除去反应期间形成的副产物。

4. 发明申请

US20130273579A1 METHOD FOR ANALYZING PROTEINS CONTRIBUTING TO AUTOIMMUNE DISEASES, AND METHOD FOR TESTING FOR SAID DISEASES 有权
标题翻译：用于分析对自身免疫性疾病作出贡献的蛋白质的方法和用于测定疾病的方法
公开(公告)号：US20130273579A1
公开(公告)日：2013-10-17
申请号：US13988269
申请日：2011-11-16
申请人： Tatsuya Sawasaki , Yaeta Endo , Tomoaki Ishigami , Ichiro Aoki
发明人： Tatsuya Sawasaki , Yaeta Endo , Tomoaki Ishigami , Ichiro Aoki
IPC分类号： G01N33/68
CPC分类号： G01N33/6869 , G01N33/564 , G01N33/6845 , G01N2800/323
摘要： Provided are a detection method for a myriad of proteins involved in an autoimmune disease with high sensitivity and high efficiency, and an analysis method for data resulting from the detection method. In order to construct the detection method and analysis method, there is provided means for comprehensively analyzing the proteins involved in an autoimmune disease by bringing a mammal-derived protein expressed in a cell-free protein synthesis system into contact with a sample derived from a patient with an autoimmune disease to detect autoantibody production, and subjecting the detected data to statistical analysis processing, and further, gene ontology analysis and/or pathway analysis.
摘要翻译：提供了涉及高灵敏度，高效率的自身免疫性疾病的无数蛋白质的检测方法以及由检测方法得到的数据的分析方法。为了构建检测方法和分析方法，提供了通过将在无细胞蛋白质合成系统中表达的哺乳动物衍生的蛋白质与来自患者的样品接触来全面分析参与自身免疫疾病的蛋白质的方法具有自身免疫疾病以检测自身抗体产生，并对检测到的数据进行统计分析处理，以及基因本体分析和/或途径分析。

5. 发明授权

US07273615B2 Reaction solution for cell-free protein synthesis, method of preparing the same, and protein synthesis method using the same 失效
标题翻译：用于无细胞蛋白质合成的反应溶液，其制备方法和使用其的蛋白质合成方法
公开(公告)号：US07273615B2
公开(公告)日：2007-09-25
申请号：US10506127
申请日：2003-02-28
申请人： Yaeta Endo , Takayasu Kawasaki , Tatsuya Sawasaki
发明人： Yaeta Endo , Takayasu Kawasaki , Tatsuya Sawasaki
IPC分类号： A61K36/00 , A61K38/00
CPC分类号： C07K16/12 , C07K16/1235 , C07K2317/622 , C12P21/00 , C12P21/02
摘要： The present invention provides a cell-free protein synthesis method characterized by a cell-free protein synthesis reaction solution that is weakly reducing and suitable for protein folding, performing a weakly reducing synthesis reaction, and preferably performing a translation reaction with the further addition of a substance catalyzing a disulfide bond exchange reaction at the beginning of the translation reaction. This method allows for proper formation of intramolecular disulfide bonds in the protein and efficiently provides protein having substantially the same function as the original protein. Specifically, it provides antibody protein that binds specifically to an antigen.
摘要翻译：本发明提供了一种无细胞蛋白质合成方法，其特征在于无细胞蛋白质合成反应溶液，其弱减少并适于蛋白质折叠，进行弱还原合成反应，优选进行平移反应，进一步加入在翻译反应开始时催化二硫键交换反应的物质。该方法允许在蛋白质中适当形成分子内二硫键，并有效地提供与原始蛋白质具有基本上相同功能的蛋白质。具体地说，它提供了与抗原特异性结合的抗体蛋白质。

6. 发明申请

US20070196812A1 Effective method of function analysis and screening of protein utilizing fluorescent light generated by cell-free protein synthesizing system 审中-公开
标题翻译：利用无细胞蛋白质合成系统产生荧光的功能分析和筛选蛋白质的有效方法
公开(公告)号：US20070196812A1
公开(公告)日：2007-08-23
申请号：US11546860
申请日：2006-10-12
申请人： Tamiyo Kobayashi , Yaeta Endo , Tatsuya Sawasaki
发明人： Tamiyo Kobayashi , Yaeta Endo , Tatsuya Sawasaki
IPC分类号： C12Q1/00
CPC分类号： G01N33/68 , G01N33/543
摘要： A method of detecting a reaction between a fluorescently labeled protein synthesized in a cell-free protein synthesizing system and a sample solution simply, in a short time and at high precision is provided. A case of detecting a binding reaction between an antibody fused with GFP and a sugar on the nanoparticle surface is explained. In a well of a microplate, a solution containing an antibody fused with GFP, and a solution containing a nanoparticle with a sugar reactive with the antibody fused with GFP adhered to a surface thereof are mixed to prepare a mixed solution A, and after the reaction, FCS measurement is performed.
摘要翻译：提供了一种在短时间和高精度下简单地检测在无细胞蛋白质合成系统中合成的荧光标记的蛋白质与样品溶液之间的反应的方法。说明检测与GFP融合的抗体与纳米粒子表面上的糖的结合反应的情况。在微孔板的孔中，混合含有与GFP融合的抗体的溶液以及含有与与其结合的GFP融合的抗体反应的糖的纳米颗粒的溶液，以制备混合溶液A，并且在反应后进行FCS测量。

7. 发明申请

US20070190579A1 Preparation method of biotinylated protein and detection method using the same 有权
标题翻译：生物素化蛋白质的制备方法及其检测方法
公开(公告)号：US20070190579A1
公开(公告)日：2007-08-16
申请号：US11643737
申请日：2006-12-21
申请人： Yaeta Endo , Tatsuya Sawasaki , Yuko Matsubara
发明人： Yaeta Endo , Tatsuya Sawasaki , Yuko Matsubara
IPC分类号： G01N33/53 , C07K14/47
CPC分类号： G01N33/532 , C07K1/13 , G01N33/543 , G01N33/58 , G01N2500/00
摘要： The present invention presents construction of a detection method requiring no step of removing free biotin during preparation of a biotinylated protein having a biotin tag, in a detection method of a substance interacting with a protein, and studied various preparation methods of the biotinylated protein. In order to solve the above-mentioned problem, the present inventor has found that in a cell-free protein synthesizing system, in particular, a wheat embryo cell-free protein synthesizing system, when biotinylation is performed during or after protein's synthesis, the biotinylation of the protein can be attained in an remarkably lower concentration of the biotin than that in the conventional biotinylation operations, and has accomplished the present invention by using the protein having the biotin tag in each detection system.
摘要翻译：本发明提出了在与蛋白质相互作用的物质的检测方法中，在制备具有生物素标签的生物素化蛋白质的过程中不需要除去游离生物素的步骤的检测方法的结构，并且研究了生物素化蛋白质的各种制备方法。为了解决上述问题，本发明人发现，在无细胞蛋白质合成系统中，特别是小麦胚细胞无蛋白质合成系统中，当在蛋白质合成期间或之后进行生物素化时，生物素化的蛋白质可以在生物素浓度显着低于常规生物素化操作中获得，并且通过在每个检测系统中使用具有生物素标签的蛋白质来完成本发明。

8. 发明申请

US20130149317A1 MALARIA VACCINE 审中-公开
公开(公告)号：US20130149317A1
公开(公告)日：2013-06-13
申请号：US13809462
申请日：2011-07-04
申请人： Takafumi Tsuboi , Motomi Torii , Tatsuya Sawasaki , Yaeta Endo
发明人： Takafumi Tsuboi , Motomi Torii , Tatsuya Sawasaki , Yaeta Endo
IPC分类号： A61K39/015 , A61K39/395
CPC分类号： A61K39/015 , A61K39/39575 , A61K2039/507 , C07K14/445 , C07K16/205 , Y02A50/412
摘要： The present invention relates to a malaria vaccine comprising:(a) a polypeptide consisting of an amino acid sequence of SEQ ID NO: 1, 2, or 3;(b) a polypeptide consisting of an amino acid sequence of SEQ ID NO: 1, 2, or 3, wherein one or more amino acids are deleted, substituted and/or added and having effect for preventing falciparum malaria; or(c) a polypeptide consisting of an amino acid sequence having 70% or more identity with an amino acid sequence of SEQ ID NO: 1, 2, or 3 and having effect for preventing falciparum malaria.
摘要翻译：本发明涉及疟疾疫苗，其包含：（a）由SEQ ID NO：1,2或3的氨基酸序列组成的多肽; （b）由SEQ ID NO：1,2或3的氨基酸序列组成的多肽，其中一个或多个氨基酸被缺失，取代和/或添加并具有预防恶性疟原虫的作用; 或（c）由与SEQ ID NO：1,2或3的氨基酸序列具有70％以上同一性并具有预防恶性疟原虫疟疾作用的氨基酸序列组成的多肽。

9. 发明申请

US20060188947A1 Reagent for producing a protein chip 审中-公开
标题翻译：用于生产蛋白质芯片的试剂
公开(公告)号：US20060188947A1
公开(公告)日：2006-08-24
申请号：US10565494
申请日：2004-08-05
申请人： Yaeta Endo , Tatsuya Sawasaki
发明人： Yaeta Endo , Tatsuya Sawasaki
IPC分类号： G01N33/53 , C12P21/06
CPC分类号： C12P21/00
摘要： An object of the present invention is to provide a reagent for protein synthesis by a simple operation which is advantageous in a cell-free protein synthesis in many samples in a test using an aimed (specific) protein or a high throughput analysis using a protein chip, etc. and also to provide a method for the synthesis of protein using the same. Further objects are to provide a protein library using the above-mentioned reagent, a kit for preparing a protein chip, a kit for testing an aimed (specific) protein, etc.
摘要翻译：本发明的目的是通过简单的操作提供用于蛋白质合成的试剂，其在使用目标（特异性）蛋白质的测试中使用蛋白质芯片的高通量分析中有利于许多样品中的无细胞蛋白质合成等等，并且还提供了使用其合成蛋白质的方法。进一步的目的是提供使用上述试剂的蛋白质文库，用于制备蛋白质芯片的试剂盒，用于测试目标（特定）蛋白质等的试剂盒。

10. 发明申请

US20060029999A1 Nucleic acid sequences having an activity of regulating translation efficiency and utilization thereof 审中-公开
标题翻译：具有调节翻译效率和利用它的活性的核酸序列
公开(公告)号：US20060029999A1
公开(公告)日：2006-02-09
申请号：US11053594
申请日：2005-02-08
申请人： Tatsuya Sawasaki , Yaeta Endo , Nami Kamura
发明人： Tatsuya Sawasaki , Yaeta Endo , Nami Kamura
IPC分类号： C12N15/867 , C07H21/04 , C12P21/06 , C07K14/47
CPC分类号： C07K14/4702 , C12N15/1051 , C12N2840/105
摘要： The present invention provides a polynucleotide comprising a nucleic acid sequence having an activity of regulating the translation efficiency of a template in a cell-free protein synthesis system and also provides a method for utilizing the same, etc. Protein synthesis is carried out by a translation template containing a polynucleotide comprising a nucleic acid sequence which is to be an object to be selected, a polyribosome fraction is prepared from the reaction solution and a nucleic acid sequence bonding to ribosome is analyzed whereupon a selection is done.
摘要翻译：本发明提供了包含具有调节无细胞蛋白质合成系统中模板的翻译效率的活性的核酸序列的多核苷酸，还提供了利用其的方法等。蛋白质合成通过翻译包含含有待选择对象的核酸序列的多核苷酸的模板，从反应溶液制备多核糖体部分，分析与核糖体结合的核酸序列，进行选择。

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