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公开(公告)号：US08318481B2
公开(公告)日：2012-11-27
申请号：US12328971
申请日：2008-12-05
申请人： Lawrence C. Chew , Stacey L. Lee
发明人： Lawrence C. Chew , Stacey L. Lee
摘要： Provided herein are improved copy number plasmids, particularly those plasmids capable of replication in a bacterial cell. The improved copy number plasmid contain a deletion, insertion, or substitution in the replication control region, particularly a Pseudomonas-specific replication control region, that results in an increase in plasmid copy number in comparison to a control plasmid. Also provided are host cells containing the improved copy number plasmids, as well as methods of using the improved copy number plasmids for the recombinant production of a protein of interest. Further provided are methods for generating plasmids with improved copy number. The methods disclosed herein involve the reiterative selection of improved copy number plasmids by the growth and selection of plasmids capable of growth under increasing selective pressure, wherein the selective pressure is applied utilizing a selection agent to which the control plasmid confers resistance.
摘要翻译： 本文提供了改进的拷贝数质粒，特别是能够在细菌细胞中复制的那些质粒。 改良的拷贝数质粒在复制控制区，特别是假单胞菌属特异性复制控制区中含有缺失，插入或取代，与对照质粒相比导致质粒拷贝数增加。 还提供了含有改进的拷贝数质粒的宿主细胞，以及使用改进的拷贝数质粒用于重组产生感兴趣的蛋白质的方法。 还提供了用于产生具有改进的拷贝数的质粒的方法。 本文公开的方法涉及通过在增加的选择压力下能够生长的质粒的生长和选择来重复选择改良的拷贝数质粒，其中使用对照质粒赋予抗性的选择剂施用选择压力。
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公开(公告)号：US20090162898A1
公开(公告)日：2009-06-25
申请号：US12328971
申请日：2008-12-05
申请人： Lawrence C. Chew , Stacey L. Lee
发明人： Lawrence C. Chew , Stacey L. Lee
摘要： Provided herein are improved copy number plasmids, particularly those plasmids capable of replication in a bacterial cell. The improved copy number plasmid contain a deletion, insertion, or substitution in the replication control region, particularly a Pseudomonas-specific replication control region, that results in an increase in plasmid copy number in comparison to a control plasmid. Also provided are host cells containing the improved copy number plasmids, as well as methods of using the improved copy number plasmids for the recombinant production of a protein of interest. Further provided are methods for generating plasmids with improved copy number. The methods disclosed herein involve the reiterative selection of improved copy number plasmids by the growth and selection of plasmids capable of growth under increasing selective pressure, wherein the selective pressure is applied utilizing a selection agent to which the control plasmid confers resistance.
摘要翻译： 本文提供了改进的拷贝数质粒，特别是能够在细菌细胞中复制的那些质粒。 改良的拷贝数质粒在复制控制区，特别是假单胞菌属特异性复制控制区中含有缺失，插入或取代，与对照质粒相比导致质粒拷贝数增加。 还提供了含有改进的拷贝数质粒的宿主细胞，以及使用改进的拷贝数质粒用于重组产生感兴趣的蛋白质的方法。 还提供了用于产生具有改进的拷贝数的质粒的方法。 本文公开的方法涉及通过在增加的选择压力下能够生长的质粒的生长和选择来重复选择改良的拷贝数质粒，其中使用对照质粒赋予抗性的选择剂施加选择压力。
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公开(公告)号：US07833752B2
公开(公告)日：2010-11-16
申请号：US12604061
申请日：2009-10-22
CPC分类号： C12N15/625 , C07K2319/02 , C07K2319/036
摘要： Compositions and methods for improving expression and/or secretion of a polypeptide of interest in a host cell are provided. Compositions including a coding sequence for a bacterial secretion signal peptide are provided. The compositions of the invention are useful for increasing accumulation of properly processed proteins in the periplasmic space of a host cell, or for increasing secretion of properly processed proteins. In particular, isolated secretion signal peptide-encoding nucleic acid molecules are provided. Additionally, amino acid sequences corresponding to the nucleic acid molecules are encompassed. The present invention provides for isolated nucleic acid molecules including nucleotide sequences encoding the amino acid sequences shown in SEQ ID NO:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, and 24, and the nucleotide sequences set forth in SEQ ID NO:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, and 23, as well as variants and fragments thereof.
摘要翻译： 提供用于改善宿主细胞中感兴趣的多肽的表达和/或分泌的组合物和方法。 提供了包括细菌分泌信号肽的编码序列的组合物。 本发明的组合物可用于增加适当加工的蛋白质在宿主细胞的周质空间中的积累，或用于增加适当加工的蛋白质的分泌。 特别地，提供了分离信号肽编码核酸分子。 另外，包含对应于核酸分子的氨基酸序列。 本发明提供了分离的核酸分子，包括编码SEQ ID NO：2,4,6,8,10,12,14,16,18,20,22和24所示的氨基酸序列的核苷酸序列， SEQ ID NO：1,3,5,7,9,11,13,15,17,19,21和23所示的核苷酸序列，以及其变体和片段。
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公开(公告)号：US09458487B2
公开(公告)日：2016-10-04
申请号：US11400840
申请日：2006-04-07
申请人： Diane M. Retallack , Charles H. Squires , David C. Watkins , Stacey L. Lee , Frank H. Gaertner , Robert Shutter
发明人： Diane M. Retallack , Charles H. Squires , David C. Watkins , Stacey L. Lee , Frank H. Gaertner , Robert Shutter
CPC分类号： C12P21/00 , C07K14/52 , C07K14/57 , C07K14/61 , C07K16/40 , C07K2317/14 , C07K2317/622 , C12N15/78 , C12P21/02 , C12Y302/01023
摘要： The invention is a process for improved production of a recombinant mammalian protein by expression in a Pseudomonad, particularly in a Pseudomonas fluorescens organism. The process improves production of mammalian proteins, particularly human or human-derived proteins, over known expression systems such as E. coli in comparable circumstances Processes for improved production of isolated mammalian, particularly human, proteins are provided.
摘要翻译： 本发明是通过在假单胞菌，特别是荧光假单胞菌生物体中表达来改进重组哺乳动物蛋白质生产的方法。 该方法在可比较的情况下，通过已知的表达系统如大肠杆菌改善了哺乳动物蛋白质，特别是人源或人源蛋白质的产生。提供了分离的哺乳动物，特别是人类蛋白质的改进生产的方法。
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公开(公告)号：US09453251B2
公开(公告)日：2016-09-27
申请号：US11038901
申请日：2005-01-18
申请人： Diane M. Retallack , Charles H. Squires , David C. Watkins , Stacey L. Lee , Frank H. Gaertner , Robert Shutter
发明人： Diane M. Retallack , Charles H. Squires , David C. Watkins , Stacey L. Lee , Frank H. Gaertner , Robert Shutter
CPC分类号： C12P21/00 , C07K14/52 , C07K14/57 , C07K14/61 , C07K16/40 , C07K2317/14 , C07K2317/622 , C12N15/78 , C12P21/02 , C12Y302/01023
摘要： The invention is a process for improved production of a recombinant mammalian protein by expression in a Pseudomonad, particularly in a Pseudomonas fluorescens organism. The process improves production of mammalian proteins, particularly human or human-derived proteins, over known expression systems such as E. coli in comparable circumstances Processes for improved production of isolated mammalian, particularly human, proteins are provided.
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公开(公告)号：US20110020868A1
公开(公告)日：2011-01-27
申请号：US12884095
申请日：2010-09-16
CPC分类号： C12N15/625 , C07K2319/02 , C07K2319/036
摘要： Compositions and methods for improving expression and/or secretion of a polypeptide of interest in a host cell are provided. Compositions including a coding sequence for a bacterial secretion signal peptide are provided. The compositions of the invention are useful for increasing accumulation of properly processed proteins in the periplasmic space of a host cell, or for increasing secretion of properly processed proteins. In particular, isolated secretion signal peptide-encoding nucleic acid molecules are provided. Additionally, amino acid sequences corresponding to the nucleic acid molecules are encompassed. The present invention provides for isolated nucleic acid molecules including nucleotide sequences encoding the amino acid sequences shown in SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, and 24, and the nucleotide sequences set forth in SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, and 23, as well as variants and fragments thereof.
摘要翻译： 提供用于改善宿主细胞中感兴趣的多肽的表达和/或分泌的组合物和方法。 提供了包括细菌分泌信号肽的编码序列的组合物。 本发明的组合物可用于增加适当加工的蛋白质在宿主细胞的周质空间中的积累，或用于增加适当加工的蛋白质的分泌。 特别地，提供了分离信号肽编码核酸分子。 另外，包含对应于核酸分子的氨基酸序列。 本发明提供了分离的核酸分子，包括编码SEQ ID NO：2,4,6,8,10,12,14,16,18,20,22和24所示的氨基酸序列的核苷酸序列， SEQ ID NO：1,3,5,7,9,11,13,15,17,19,21和23所示的核苷酸序列，以及其变体和片段。
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