The present invention relates in part to nucleic acids encoding proteins, nucleic acids containing non-canonical nucleotides, therapeutics comprising nucleic acids, methods, kits, and devices for inducing cells to express proteins, methods, kits, and devices for transfecting, gene editing, and reprogramming cells, and cells, organisms, and therapeutics produced using these methods, kits, and devices. Methods for inducing cells to express proteins and for reprogramming and gene-editing cells using RNA are disclosed. Methods for producing cells from patient samples, cells produced using these methods, and therapeutics comprising cells produced using these methods are also disclosed.

The present disclosure provides, among other things, vectors for expression of ENPP1 or ENPP3 in vivo and methods for the treatment of diseases of calcification and ossification in a subject.

Provided herein are novel blocking chimeric antigen receptors (“bCARs”) and immune cells (e.g., effector and regulatory immune cells) that express such bCARs. Such blocking CARs prevent undesired activation of the immune cells, particularly undesired activation of the immune cells against normal tissue in therapeutic applications. Thus, such bCARs advantageously allow for selective immune cell activation only upon interaction with specific target cells (e.g., tumor cell).

Various examples according to the present disclosure provide a fermentation method. The fermentation method includes producing at least about 10 g/L of a bioproduct and one or more heterologous polypeptides by fermenting a medium using an engineered microorganism. About 2 wt % to about 100 wt % of the one or more heterologous polypeptides are encapsulated intercellularly in the engineered microorganism. The method further includes isolating the engineered microorganism including the encapsulated one or more heterologous polypeptides. About 50 wt % to about 100 wt % of the one or more heterologous polypeptides retain functionality following isolation of the engineered microorganism.

Various examples according to the present disclosure provide a fermentation method. The fermentation method includes producing at least about 10 g/L of a bioproduct and one or more heterologous polypeptides by fermenting a medium using an engineered microorganism. About 2 wt % to about 100 wt % of the one or more heterologous polypeptides are encapsulated intercellularly in the engineered microorganism. The method further includes isolating the engineered microorganism including the encapsulated one or more heterologous polypeptides. About 50 wt % to about 100 wt % of the one or more heterologous polypeptides retain functionality following isolation of the engineered microorganism.

The present invention provides a nucleic acid construct comprising the following structure: A-X-B in which A is nucleic acid sequence encoding a first polypeptide which comprises a first signal peptide; B is nucleic acid sequence encoding a second polypeptide which comprises a second signal peptide and X is a nucleic acid sequence which encodes a cleavage site, wherein the first signal peptide or the second signal peptide comprises one or more mutation(s) such that it has fewer hydrophobic amino acids.

The present invention provides a nucleic acid construct comprising the following structure: A-X-B in which A is nucleic acid sequence encoding a first polypeptide which comprises a first signal peptide; B is nucleic acid sequence encoding a second polypeptide which comprises a second signal peptide and X is a nucleic acid sequence which encodes a cleavage site, wherein the first signal peptide or the second signal peptide comprises one or more mutation(s) such that it has fewer hydrophobic amino acids.

The invention relates to methods, uses, systems, arrays, engineered nucleotide sequences and vectors for inhibiting bacterial population growth or for altering the relative ratio of sub-populations of first and second bacteria in a mixed population of bacteria. The invention is particularly useful, for example, for treatment of microbes such as for environmental, medical, food and beverage use. The invention relates inter alia to methods of controlling microbiologically influenced corrosion (MIC) or biofouling of a substrate or fluid in an industrial or domestic system.

The invention relates to methods, uses, systems, arrays, engineered nucleotide sequences and vectors for inhibiting bacterial population growth or for altering the relative ratio of sub-populations of first and second bacteria in a mixed population of bacteria. The invention is particularly useful, for example, for treatment of microbes such as for environmental, medical, food and beverage use. The invention relates inter alia to methods of controlling microbiologically influenced corrosion (MIC) or biofouling of a substrate or fluid in an industrial or domestic system.

Compositions and methods are described for the delivery of recombinant human iduronate-2-sulfatase (IDS) produced by human neuronal or glial cells to the cerebrospinal fluid of the central nervous system (CNS) of a human subject diagnosed with mucopolysaccharidosis II (MPS II).