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公开(公告)号：US08288127B2
公开(公告)日：2012-10-16
申请号：US12512930
申请日：2009-07-30
CPC分类号： C12N9/0028 , C12N9/88 , C12N15/52 , C12N15/78 , C12P21/00 , C12P21/02 , C12Y305/05001
摘要： The present invention provides an improved expression system for the production of recombinant polypeptides utilizing auxotrophic selectable markers. In addition, the present invention provides improved recombinant protein production in host cells through the improved regulation of expression.
摘要翻译： 本发明提供了利用营养缺陷型选择标记物生产重组多肽的改进的表达系统。 此外，本发明通过改进的表达调节来提供在宿主细胞中改进的重组蛋白质产生。
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公开(公告)号：US20090325230A1
公开(公告)日：2009-12-31
申请号：US12512930
申请日：2009-07-30
IPC分类号： C12P21/00
CPC分类号： C12N9/0028 , C12N9/88 , C12N15/52 , C12N15/78 , C12P21/00 , C12P21/02 , C12Y305/05001
摘要： The present invention provides an improved expression system for the production of recombinant polypeptides utilizing auxotrophic selectable markers. In addition, the present invention provides improved recombinant protein production in host cells through the improved regulation of expression.
摘要翻译： 本发明提供了利用营养缺陷型选择标记物生产重组多肽的改进的表达系统。 此外，本发明通过改进的表达调节来提供在宿主细胞中改进的重组蛋白质产生。
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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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4.发明申请公开(公告)号：US20160208268A1
公开(公告)日：2016-07-21
申请号：US14974897
申请日：2015-12-18
IPC分类号： C12N15/65
摘要： Provided herein are methods and compositions for expression of a nucleic acid construct comprising nucleic acids encoding a) a recombinant polypeptide, and b) a prototrophy-restoring enzyme in a host cell that is auxotrophic for at least one metabolite. In various embodiments, the host cell is auxotrophic for a nitrogenous base compound or an amino acid. The invention involves introducing an analogue into the growth media for the host cell such that the analogue is incorporated into the recombinant polypeptide or a nucleic acid coding sequence thereof. In various embodiments, the compositions and methods disclosed herein result in improved recombinant protein expression compared to expression of recombinant protein in an antibiotic selection system, or compared to expression of the recombinant protein in an expression system that lacks a metabolite analogue.
摘要翻译： 本文提供了用于表达包含编码a）重组多肽的核酸的核酸构建体的方法和组合物，以及b）对于至少一种代谢产物是营养缺陷型的宿主细胞中的原始营养恢复酶。 在各种实施方案中，宿主细胞对于含氮碱性化合物或氨基酸是营养缺陷型的。 本发明涉及将类似物引入宿主细胞的生长培养基中，使得类似物掺入重组多肽或其核酸编码序列中。 在各种实施方案中，与抗生素选择系统中的重组蛋白的表达相比，本文公开的组合物和方法导致改善的重组蛋白表达，或与缺乏代谢物类似物的表达系统中重组蛋白的表达进行比较。
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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：US09708616B2
公开(公告)日：2017-07-18
申请号：US14974897
申请日：2015-12-18
摘要： Provided herein are methods and compositions for expression of a nucleic acid construct comprising nucleic acids encoding a) a recombinant polypeptide, and b) a prototrophy-restoring enzyme in a host cell that is auxotrophic for at least one metabolite. In various embodiments, the host cell is auxotrophic for a nitrogenous base compound or an amino acid. The invention involves introducing an analog into the growth media for the host cell such that the analog is incorporated into the recombinant polypeptide or a nucleic acid coding sequence thereof. In various embodiments, the compositions and methods disclosed herein result in improved recombinant protein expression compared to expression of recombinant protein in an antibiotic selection system, or compared to expression of the recombinant protein in an expression system that lacks a metabolite analog.
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7.发明授权公开(公告)号：US09243253B2
公开(公告)日：2016-01-26
申请号：US13319844
申请日：2010-05-10
摘要： Provided herein are methods and compositions for expression of a nucleic acid construct comprising nucleic acids encoding a) a recombinant polypeptide, and b) a prototrophy-restoring enzyme in a host cell that is auxotrophic for at least one metabolite. In various embodiments, the host cell is auxotrophic for a nitrogenous base compound or an amino acid. The invention involves introducing an analogue into the growth media for the host cell such that the analogue is incorporated into the recombinant polypeptide or a nucleic acid coding sequence thereof. In various embodiments, the compositions and methods disclosed herein result in improved recombinant protein expression compared to expression of recombinant protein in an antibiotic selection system, or compared to expression of the recombinant protein in an expression system that lacks a metabolite analogue.
摘要翻译： 本文提供了用于表达包含编码a）重组多肽的核酸的核酸构建体的方法和组合物，以及b）对于至少一种代谢产物是营养缺陷型的宿主细胞中的原始营养恢复酶。 在各种实施方案中，宿主细胞对于含氮碱性化合物或氨基酸是营养缺陷型的。 本发明涉及将类似物引入宿主细胞的生长培养基中，使得类似物掺入重组多肽或其核酸编码序列中。 在各种实施方案中，与抗生素选择系统中的重组蛋白的表达相比，本文公开的组合物和方法导致改善的重组蛋白表达，或与缺乏代谢物类似物的表达系统中重组蛋白的表达进行比较。
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公开(公告)号：US07745167B2
公开(公告)日：2010-06-29
申请号：US11528872
申请日：2006-09-28
CPC分类号： G01N21/474 , G01N21/51
摘要： A method for monitoring fluid media, such as a dynamic biological system, in a biological reactor containing developing culture fluid media. The method includes the step of directing light into the fluid media by way of one or more optical fibers to produce an illuminated fluid media and then measuring the intensity of light reflected from the illuminated fluid media by way of one or more optical fibers, the optical fibers being partitioned from the fluid media by a transparent window having inner and outer surfaces; the fibers having ends terminating adjacent to and confronting the inner surface of the window and extending in a direction away from the window, the corresponding ends of the fibers being radially and circumferentially spaced from one another, the corresponding ends of the fibers having converging and intersecting longitudinal projections therefrom, the intersecting longitudinal projections from the fibers being entirely within the window. Additionally, a fiber-optic probe is discussed.
摘要翻译： 一种用于在含有显影培养液介质的生物反应器中监测流体介质如动态生物系统的方法。 该方法包括通过一根或多根光纤将光引导到流体介质中以产生照明的流体介质，然后通过一根或多根光纤测量从被照射的流体介质反射的光的强度的步骤，该光学 纤维通过具有内表面和外表面的透明窗口与流体介质隔开; 纤维的端部邻近并面向窗口的内表面并且沿远离窗口的方向延伸，纤维的相应端部彼此径向和周向间隔开，纤维的相应端部具有会聚和相交 来自纤维的交叉纵向突起完全在窗内。 另外，讨论了光纤探针。
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