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    Production of recombinant proteins utilizing non-antibiotic selection methods and the incorporation of non-natural amino acids therein
    2.
    发明授权
    Production of recombinant proteins utilizing non-antibiotic selection methods and the incorporation of non-natural amino acids therein 有权
    使用非抗生素选择方法生产重组蛋白，并在其中掺入非天然氨基酸

    公开(公告)号：US09243253B2

    公开(公告)日：2016-01-26

    申请号：US13319844

    申请日：2010-05-10

    申请人： Diane M. Retallack ,  Lawrence C. Chew ,  Charles H. Squires

    发明人： Diane M. Retallack ,  Lawrence C. Chew ,  Charles H. Squires

    IPC分类号： C12N15/00 ,  C12N15/65 ,  C12P21/02 ,  C12N15/78 ,  C12N15/63

    CPC分类号： C12N15/65 ,  C12N15/63 ,  C12N15/78 ,  C12P21/02

    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。

    PROCESS FOR IMPROVED PROTEIN EXPRESSION BY STRAIN ENGINEERING
    3.
    发明申请
    PROCESS FOR IMPROVED PROTEIN EXPRESSION BY STRAIN ENGINEERING 有权
    通过应变工程改进蛋白质表达的方法

    公开(公告)号：US20140162279A1

    公开(公告)日：2014-06-12

    申请号：US14071273

    申请日：2013-11-04

    申请人： PFENEX INC.

    发明人： THOMAS M. RAMSEIER ,  HONGFAN JIN ,  CHARLES H. SQUIRES

    IPC分类号： C12N15/63

    CPC分类号： C12N15/63 ,  C12N15/70 ,  C12N15/74 ,  C12N15/78 ,  C12P21/02

    摘要： This invention is a process for improving the production levels of recombinant proteins or peptides or improving the level of active recombinant proteins or peptides expressed in host cells. The invention is a process of comparing two genetic profiles of a cell that expresses a recombinant protein and modifying the cell to change the expression of a gene product that is upregulated in response to the recombinant protein expression. The process can improve protein production or can improve protein quality, for example, by increasing solubility of a recombinant protein.

    摘要翻译： 本发明是改善重组蛋白或肽的生产水平或提高在宿主细胞中表达的活性重组蛋白或多肽的水平的方法。 本发明是比较表达重组蛋白的细胞的两种遗传图谱并修饰细胞以改变响应于重组蛋白表达上调的基因产物的表达的过程。 该方法可以改善蛋白质产生或者可以通过增加重组蛋白质的溶解度来改善蛋白质质量。

    Mannitol induced promoter systems in bacterial host cells
    5.
    发明授权
    Mannitol induced promoter systems in bacterial host cells 有权
    甘露醇诱导细菌宿主细胞中的启动子系统

    公开(公告)号：US08017355B2

    公开(公告)日：2011-09-13

    申请号：US12330723

    申请日：2008-12-09

    申请人： J. Carrie Schneider ,  Bettina Rosner

    发明人： J. Carrie Schneider ,  Bettina Rosner

    IPC分类号： C12P21/00 ,  C12N1/21 ,  C07H21/04

    CPC分类号： C07K14/21 ,  C12N15/78 ,  C12P21/02

    摘要： The present invention provides methods for producing recombinant peptides in a bacterial host utilizing a mannitol, arabitol, glucitol, or glycerol-inducible promoter, wherein the host bacterial cell that produces the peptide has been rendered incapable of degrading or metabolizing mannitol, arabitol, or glucitol, or derivatives or analogues thereof. The present invention provides bacterial cells that have been genetically altered to inhibit the metabolism or degradation of mannitol, glucitol, or arabitol, or derivatives or analogues thereof. The present invention utilizes mannitol, arabitol, glucitol, or glycerol to induce expression of a target polypeptide from an inducible promoter, allowing for the use of an inexpensive and stable carbon source inducer in the fermentation processes for the production of recombinant peptides.

    摘要翻译： 本发明提供了利用甘露醇，阿糖醇，葡萄糖醇或甘油诱导型启动子在细菌宿主中产生重组肽的方法，其中产生该肽的宿主细菌细胞已经不能降解或代谢甘露醇，阿糖醇或葡萄糖醇 ，或其衍生物或类似物。 本发明提供已被遗传改变以抑制甘露醇，葡萄糖醇或阿拉伯糖醇或其衍生物或类似物的代谢或降解的细菌细胞。 本发明利用甘露醇，阿糖醇，葡萄糖醇或甘油从诱导型启动子诱导靶多肽的表达，允许在用于生产重组肽的发酵过程中使用便宜且稳定的碳源诱导剂。

    Method for rapidly screening microbial hosts to identify certain strains with improved yield and/or quality in the expression of heterologous proteins
    7.
    发明授权
    Method for rapidly screening microbial hosts to identify certain strains with improved yield and/or quality in the expression of heterologous proteins 有权
    用于快速筛选微生物宿主以鉴定具有提高异源蛋白质表达的产量和/或质量的某些菌株的方法

    公开(公告)号：US09580719B2

    公开(公告)日：2017-02-28

    申请号：US12610207

    申请日：2009-10-30

    申请人： Diane M. Retallack ,  Charles H. Squires ,  Thomas M. Ramseier ,  Russell J. Coleman ,  Jane C. Schneider ,  Charles D. Hershberger

    发明人： Diane M. Retallack ,  Charles H. Squires ,  Thomas M. Ramseier ,  Russell J. Coleman ,  Jane C. Schneider ,  Charles D. Hershberger

    IPC分类号： C12N15/78

    CPC分类号： C12N15/1037 ,  C12N15/78

    摘要： The present invention provides an array for rapidly identifying a host cell population capable of producing a heterologous protein with improved yield and/or quality. The array comprises one or more host cell populations that have been genetically modified to increase the expression of one or more target genes involved in protein production, decrease the expression of one or more target genes involved in protein degradation, or both. One or more of the strains in the array may express the heterologous protein of interest in a periplasm compartment or may secrete the heterologous protein extracellularly through an outer cell wall. The strain arrays are useful for screening for improved expression of any protein of interest including therapeutic proteins, hormones, growth factors, extracellular receptors or ligands, proteases, kinases, blood proteins, chemokines, cytokines, antibodies and the like.

    摘要翻译： 本发明提供了一种用于快速鉴定能够产生具有改善的产量和/或质量的异源蛋白质的宿主细胞群的阵列。 该阵列包含已被遗传修饰的一个或多个宿主细胞群，以增加参与蛋白质产生的一个或多个靶基因的表达，减少参与蛋白质降解的一个或多个靶基因的表达，或两者。 阵列中的一个或多个菌株可以在周质隔室中表达感兴趣的异源蛋白质，或者通过外细胞壁在细胞外分泌异源蛋白质。 菌株阵列可用于筛选任何目的蛋白质的表达，包括治疗性蛋白质，激素，生长因子，细胞外受体或配体，蛋白酶，激酶，血液蛋白，趋化因子，细胞因子，抗体等。

    High copy number self-replicating plasmids in pseudomonas
    10.
    发明授权
    High copy number self-replicating plasmids in pseudomonas 失效
    假单胞菌中高拷贝数自复制质粒

    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

    IPC分类号： C12N15/63 ,  C12N1/21 ,  C07K14/00

    CPC分类号： C12N15/69 ,  C12N15/78

    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。