Described herein are method for generating a vector for editing a cell. The method comprises ligating into a vector that encodes a portion of a gRNA a cassette comprising at least one editing cassette, a promoter, and a gene encoding another portion of the gRNA. Upon ligation, the portion of the gRNA from the editing cassette and the other portion of the gRNA are ligated and form a functional gRNA.

A method for expression of transcribable unit(s) in a target cell is provided. The method comprises the steps of: a) providing a target cell expressing a site-specific recombinase, b) providing a DNA vector characterized by a 5 to 3 vector sequence orientation. The DNA vector comprises a plurality of recombination units, wherein a single recombination unit comprises at least one transcribable unit and a first type and a second type of target site that are recognizable by the site-specific recombinase. Recombination can only occur between two target sites of the same type and the first type of target site is located at the 5 start of the recombination unit and the second type of target site is located at the 3 end of the recombination unit. For all recombination units comprised within the DNA vector, the orientation of all of the first type of target sites are the same, and the orientation of all of the second type of target sites are the same. Step c) comprises introducing the DNA vector into the target cell.

The present disclosure provides compositions, systems, and methods for pooled screening of perturbations correlating to a phenotype. The present disclosure also provides for methods of in vivo perturbation screening. The present disclosure also provides for high-throughput sample multiplexing. The present disclosure also provides for methods to determine the clonal effects associated with millions of genetic perturbation combinations using standard pooled screening reagents.

Methods are disclosed herein for efficiently generating human induced pluripotent stem cells (iPSC) containing a nucleic acid including a doxycycline promoter operably linked to a nucleic acid encoding Cas9. These methods include transfecting a human somatic cell with a nucleic acid molecule comprising a doxycycline promoter operably linked to a nucleic acid encoding a Cas9, and constitutive promoter operably linked to a tetracycline responsive element and inducing the somatic cell to form an iPSC, thereby producing an iPSC that can undergo CRISPR/Cas9-mediated recombination at a high efficiency. The human iPSC, or a cell differentiated therefrom, is cultured in the presence of doxycycline to induce expression of the Cas9. These cells can then be used to target in any gene of interest by introducing nucleic acids encoding sgRNAs. Induced pluripotent stem cells produced by these methods are also disclosed.

Provided herein are non-naturally occurring self-assembling polypeptides for transferring nucleic acids and/or proteins to a cell, pharmaceutical compositions comprising such polypeptides, and methods for treatment comprising use of such compositions. The methods for producing polypeptide compositions may include combining in a solution, unassembled recombinant GAG-like proteins, nucleic acids and/or proteins in low salt conditions; and increasing the ionic strength of the solution.

The present disclosure relates generally to methods for ameliorating or treating acute myeloid leukemia (AML). In particular, the present technology relates to administering a therapeutically effective amount of one or more compositions that inhibit the vitamin B6 pathway to a subject diagnosed with, or at risk for AML.

Provided is a pig genome-wide specific sgRNA library, a preparation method therefor, and an application thereof. The sgRNA is targeted at a pig genome-wide protein-coding gene, lincRNA and/or miRNA. Specifically, an sgRNA construct has the following structure: AL-N20-AR, wherein AL is the left homology arm sequence located at the upstream of the coding sequence of a pig specific SgRNA, N20 is the coding sequence of the pig specific SgRNA, and AR is a right homology arm sequence located at the downstream of the coding sequence of the pig specific sgRNA. The sgRNA library can be used for screening functional genes of a pig or for preparing a kit.

The invention provides cells and animals, as well as methods of producing cells and animals, that express at least one interfering RNA molecule to regulate the expression of a specific gene or family of genes. The invention further provides novel iRNA molecules, as well as DNA templates for producing iRNA molecules.

Cas-protein-ready tau biosensor cells, CRISPR/Cas synergistic activation mediator (SAM)-ready tau biosensor cells, and methods of making and using such cells to screen for genetic modifiers of tau seeding or aggregation are provided. Reagents and methods for sensitizing such cells to tau seeding activity or tau aggregation or for causing tau aggregation are also provided.

Described herein are methods and vectors for rational, multiplexed manipulation of chromosomes within open reading frames (e.g., in protein libraries) or any segment of a chromosome in a cell or population of cells, in which various CRISPR systems are used.