The invention involves inducing 3-50 or more mutations (e.g., any whole number between 3 and 50 of mutations, with it noted that in some embodiments there can be up to 16 different RNA(s), e.g., sgRNAs each having its own a promoter, in a vector, such as AAV, and that when each sgRNA does not have its own promoter, there can be twice to thrice that amount of different RNA(s), e.g., sgRNAs, e.g., 32 or even 48 different guides delivered by one vector) in transgenic Cas9 eukaryotes to model genetic disease, e.g. cancer. The invention comprehends testing putative treatments with such models, e.g., testing putative chemical compounds that may be pharmaceutically relevant for treatment or gene therapy that may be relevant for treatment, or combinations thereof. The invention allows for the study of genetic diseases and putative treatments to better understand and alleviate a genetic disease or a condition, e.g., cancer.

Described are receptor ligands of transient receptor potential cation channel subfamily V member 4 (TRPV4), pharmaceutical compositions including the compounds, and methods of using the compounds and compositions for treating ocular disorders.

Provided are improved compositions and methods for achieving gene therapy in hematopoietic cells and hematopoietic precursor cells, including erythrocytes, erythroid progenitors, and embryonic stem cells. Also provided are improved gene therapy methods for treating hematopoietic-related disorders. Retroviral gene therapy vectors that are optimized for erythroid specific expression and treatment of hemoglobinopathic conditions are disclosed.

Provided is a temporary treatment medium for inhibiting embryo developmental arrest due to manipulation of an embryo or the like. A temporary treatment medium according to the present invention reduces damage to an in-vitro culture due to manipulation thereof, said in-vitro culture containing one of pluripotent stem cells, reproductive cells, a fertilized egg, and an embryo or any combination thereof. For this purpose, the temporary treatment medium contains a cytoskeleton regulator and/or an apoptosis inhibitor. The cytoskeleton regulator and/or the apoptosis inhibitor is preferably an Rho kinase inhibitor. Specifically, a Rock inhibitor can be used as the Rho kinase inhibitor. The Rock inhibitor is, for example, Y-27632. The temporary treatment medium is used for treating an in-vitro culture for a specific period before and/or after manipulation involving damage thereto.

The invention provides novel engineered CRISPR/Cas effector enzymes, such as Cas13 (e.g., Cas13d, Cas13e, or Cas13f) that substantially maintain guide-sequence-specific endonuclease activity and substantially lack guide-sequence-independent collateral endonuclease activity compared to the corresponding wild-type Cas. Also provided are polynucleotides encoding the same, vectors or host cells comprising the polynucleotides or engineered Cas, and method of use, such as in RNA-based target gene transcript knock down.

In some aspects, the present invention provides chimeric transferrin receptor (TfR) polynucleotides and polypeptides. In other aspects, this invention provides chimeric TfR transgenic animal models and methods of using the animal models to identify therapeutics that can cross the blood-brain barrier.

A primatized rodent or swine, and methods of making and using the primatized rodent or swine, are provided.

Methods and compositions are provided for making non-human animals with reduced tolerance of a foreign antigen of interest and making antigen-binding proteins against that foreign antigen of interest using such animals. The methods and compositions employ CRISPR/Cas9 systems using multiple guide RNAs to reduce or eliminate expression of a self-antigen homologous to or sharing an epitope of interest with the foreign antigen of interest or to reduce or eliminate expression of an epitope on the self-antigen that is shared with the foreign antigen of interest.

The invention provides double knockout transgenic pigs (GT/CMAH-KO pigs) lacking expression of any functional αGAL and CMAH. Double knockout GT/CMAH-KO transgenic organs, tissues and cells are also provided. Methods of making and using the GT/CMAH-KO pigs and tissue are also provided.

Disclosed are a therapeutic agent for muscular dystrophy employing pluripotent stem cells obtained from dental pulp and a method for preparation thereof. The therapeutic agent for muscular dystrophy comprises pluripotent stem cell-enriched human dental pulp-derived cells as the active ingredient, and is prepared by a method of preparation comprising the steps of: (a) adding dental pulp-derived cells contained in a dental pulp suspension, in a culture vessel containing feeder cells whose proliferative ability is suppressed, onto a membrane having micropores that can block feeder cells from passing therethrough and supported within the vessel in a manner that avoids direct contact of the lower side face thereof with the feeder cells, and culturing the dental pulp-derived cells on the membrane while preventing direct contact with the feeder cells, and (b) recovering the cells having grown on the membrane as the pluripotent stem cell-enriched human dental pulp-derived cells.