The present disclosure relates to automated multi-module instruments, compositions and methods for performing nucleic acid-guided nuclease editing; specifically, methods, instruments, systems, and nucleic acids synthetic cassettes that improve the efficiency of gene editing using CRISPR enzymes in diploid cells—either on both chromosomes or selectively in one chromosome—without incurring loss of heterozygosity (LOH) and its often deleterious effects.

Provided herein are methods and compositions for modifying cells. Provided herein are methods and compositions for modifying an organism of a cell. Provided herein are methods and compositions for introducing polynucleotides and/or polypeptides into a nucleus of a cell.

The disclosure relates to a plurality of cells, compositions and methods for identifying modulators of a target protein. The cells, compositions and methods comprise a (i) a target domain gene (ii) an intracellular chimeric G-protein alpha subunit comprising an endogenous G-protein alpha subunit with a humanized C-terminus; and (iii) an inducible reporter, wherein the expression of the reporter is dependent on the activation of the target domain encoded by target domain gene, and wherein the target domain gene comprises a barcode. The disclosure further relates to a host cell comprising a plurality of exogenous landing pads integrated in the host cell's genome, wherein each exogenous landing pad is integrated at a safe harbor genome loci in the host cell's genome.

Methods for modification of target nucleic acids. The method involves a construct in which guide RNA is covalently linked to donor RNA (fusion NA) to be introduced into the target nucleic acid by homologous recombination and is based on the introduction of a nuclease, e.g. CRISPR or TALEN, into the cell containing the target nucleic acid. The fusion NA may be introduced as a DNA vector.

The present disclosure provides compositions and methods to increase the percentage of edited cells in a cell population when employing nucleic-acid guided editing, as well as automated multi-module instruments for performing these methods. Specifically, the disclosure relates to methods, compositions, modules and automated multi-module cell processing instruments that increase the efficiency of nucleic acid-guided editing in a cell population using a nucleic acid nuclease (i.e., an RNA-guided nuclease or “RGN”)/single-strand binding protein (“SSB”) fusion system. The system leverages a single-strand binding protein (SSP) and single-strand DNA annealing protein (“SSAP”) interactions to drive enhanced recruitment.

Methods for modification of target nucleic acids. The method involves a construct in which guide RNA is covalently linked to donor RNA (fusion NA) to be introduced into the target nucleic acid by homologous recombination and is based on the introduction of a nuclease, e.g. CRISPR or TALEN, into the cell containing the target nucleic acid. The fusion NA may be introduced as a DNA vector.

Disclosed herein are modified strains for improving secretion of recombinantly expressed products secreted from a host organism with improved growth and productivity characteristics, as well as methods of using the modified strains.

The present disclosure relates to compositions and methods of generating circular-permuted nucleic acids for direct use in homology directed genome editing. The method also allows the joining of a large number of DNA fragments, in a deterministic fashion. It can be used to rapidly generate nucleic acid libraries that can be directly used in a variety of applications without further cloning steps that include, for example, genome editing and pathway assembly. Kits for performing the method are also disclosed.

A novel gene targeting method and a nucleotide construct for the method. The method integrates a nucleotide construct containing an interference gene in an effective gene targeting region independent of the gene by homologous recombination, thereby improving the targeting efficiency of the gene. The present invention also provides a gene targeting system for gene expression regulation and gene disruption.

Methods for modification of target nucleic acids. The method involves a construct in which guide RNA is covalently linked to donor RNA (fusion NA) to be introduced into the target nucleic acid by homologous recombination and is based on the introduction of a nuclease, e.g. CRISPR or TALEN, into the cell containing the target nucleic acid. The fusion NA may be introduced as a DNA vector.