The present invention provides for a mechanism to reduce glycerol production and increase nitrogen utilization and ethanol production of recombinant microorganisms. One aspect of this invention relates to strains of S. cerevisiae with reduced glycerol productivity that get a kinetic benefit from higher nitrogen concentration without sacrificing ethanol yield. A second aspect of the invention relates to metabolic modifications resulting in altered transport and/or intracellular metabolism of nitrogen sources present in corn mash.

The present invention provides methods and compositions for stable genetic modification of cultured mammalian cells. The genetic modifications can be used to produce cultured mammalian cells for therapeutic or diagnostic purposes.

The present disclosure reports that a calcium-dependent serine protease, complement component is (C1s) has been identified as a protease responsive for cleavage of exogenous polypeptides expressed in mammalian cell lines such as CHO cells. These CHO cell lines provide for increased yield of antigenically correct, uncleaved exogenous polypeptide as compared to unmodified CHO cells expressing the active C1s protease. C1s-deficient cell lines and methods for use of same for producing exogenous polypeptides, e.g., human immunodeficiency virus (HIV) envelope glycoprotein polypeptides, such as, gp120 or human Factor VIII are provided.

The present disclosure relates to providing an engineered CHO cell line wherein the two essential metabolic genes are knocked out. Particularly, the present invention relates to a double knockout CHO cell line (DHFR−/− and GS−/−) with disrupted Dihydrofolate Reductase (DHFR) and Glutamine Synthetase (GS) genomic loci. The double knockout CHO cell line (DHFR−/− and GS−/−) being suitable for expression of monoclonal antibodies, dimeric therapeutic proteins, Fab, single chain fragments, or the like. The present disclosure also provides method of generation of a double knock out CHO cell line (DHFR−/− and GS−/−) using gene selection and/or manipulating techniques such as CRISPR/Cas9 system, Zinc Finger Nuclease, TALEN, or the like. The present disclosure further provides method of selection of clones and production of therapeutic proteins of interest with increased titre.

Disclosed herein are methods of increasing nitrogen fixation in a non-leguminous plant. The methods can comprise exposing the plant to a plurality of bacteria. Each member of the plurality comprises one or more genetic variations introduced into one or more genes or non-coding polynucleotides of the bacteria's nitrogen fixation or assimilation genetic regulatory network, such that the bacteria are capable of fixing atmospheric nitrogen in the presence of exogenous nitrogen. The bacteria are not intergeneric microorganisms. Additionally, the bacteria, in planta, produce 1% or more of the fixed nitrogen in the plant.

The present invention provides polynucleotide vectors for high expression of heterologous genes. Some vectors further comprise novel transposons and transposases that further improve expression. Further disclosed are vectors that can be used in a gene transfer system for stably introducing nucleic acids into the DNA of a cell. The gene transfer systems can be used in methods, for example, gene expression, bioprocessing, gene therapy, insertional mutagenesis, or gene discovery

Provided are compositions comprising polynucleotides encoding glutamine synthetase (GS) polypeptides having improved properties, such as increased enzymatic activity and/or increased thermostability. Also provided are recombinant DNA constructs, plants, plant cells, seed, grain comprising the polynucleotides. Additionally, various methods of employing the polynucleotides in plants, such as methods for increasing GS activity in a plant, methods for increase seed protein content in a seed of a plant, and methods for increasing yield of a plant, are also provided herein.

The present invention relates to the use of glutamine synthetase as a protein therapy (such as enzyme replacement protein therapy) for the treatment of hyperammonemia. In particular the invention relates to the systemic administration of glutamine synthetase. The glutamine synthetase may be provided in conjugated or fusion form, to increase its half-life in the circulation. Also provided is a pharmaceutical composition comprising glutamine synthetase. The invention also relates to the uses, methods and compositions involving a combination of the glutamine synthetase protein and an ammonia lowering agent, such as a nitrogen scavenger.

Disclosed herein are methods of manufacturing therapeutic proteins.

The present invention provides methods and compositions for stable genetic modification of cultured mammalian cells. The genetic modifications can be used to produce cultured mammalian cells for therapeutic or diagnostic purposes.