The disclosure discloses an in-vivo continuous directed evolution system and application thereof, and belongs to the fields of gene engineering and enzyme engineering. The system includes Escherichia coli host bacteria carrying a random mutation module mutagenesis plasmid, a programmed death module toxin-antitoxin system and a target gene expression module target plasmid. The modules are coupled with one another, and target genes are subjected to multiple rounds of continuous mutation by virtue of the random mutation module mutagenesis plasmid in the system, so that the mutation rate of the target genes is further increased, and ultimately, efficient evolution and screening of the target genes in the host bacteria are realized. According to the system, mutations are accurately positioned on the target genes, random mutations in non-target gene regions are reduced, and the system has good practical value and can be applied to directed evolution of various different functional proteins.

The present disclosure provides, among other things, a way to amplify and sequence target sequences in a low-input sample. In some embodiments, the method comprises ligating a double-stranded adaptor onto a population of fragments to produce tagged fragments, and linearly amplifying the tagged fragments.

The disclosure relates to methods for the screening, identification, and/or application of one or more microorganisms of use in imparting one or more beneficial properties to one or more plants.

The present disclosure provides methods and systems for identification of genomic regions for therapeutic targeting. A method for identifying one or more genomic regions for therapeutic targeting, which may facilitate re-programming of a cell from one phenotypic state to another, may comprise: providing single-cell RNA-seq data for a plurality of diseased cells and a plurality of normal cells of a cell type; mapping the single-cell RNA-seq data for the plurality of diseased cells and the plurality of normal cells into a latent space corresponding to a plurality of phenotypic states of the cell type; identifying, based at least in part on a topology of the latent space, the one or more genomic regions for therapeutic targeting; and electronically outputting the one or more genomic regions for therapeutic targeting.

The invention provides methods and means for specifically altering the DNA sequence in a genome, in particular for genome editing by deleting or replacing a sequence of interest. Advantageously, the invention uses two non-identical sequences naturally occurring in a genome as target sites two which DNA-recombining enzymes are generated. The invention is in particular useful for medicine, in particular to repair a mutation in a genome or to delete predefined genetic material from cells or tissue and to cure diseases. An advantage of the invention is that it allows precise site directed altering of DNA without engaging host DNA repair pathways and thereby works without inducing random insertions and deletions (in-dels).

Provided herein are polypeptides that bind to a transferrin receptor, methods of generating such polypeptides, and methods of using the polypeptides to target a composition to a transferrin receptor-expressing cell.

Presented herein is a high-throughput (HTP) genomic engineering platform for improving the production of therapeutic proteins in Chinese hamster ovary (CHO) cells. The disclosed HTP genomic engineering platform is computationally driven and integrates molecular biology, automation, and advanced machine learning protocols. The platform utilizes a unique suite of HTP genetic engineering tools to explore the genomic landscape associated with therapeutic protein production pathways, in order to unravel the biological drivers and disentangle the uncharacterized genetic architecture responsible for optimizing therapeutic protein production in CHO cells.

The present disclosure provides adenine base editors (ABEs) that are variants of known adenine base editors. The adenosine deaminase domain of a known ABE was modified to produce adenosine deaminase variants. The deaminase variants provided herein have broader compatibility with diverse napDNAbp domains, such as Cas homologs, for base editing applications. The ABEs provided herein comprise a deaminase variant and a napDNAbp domain. The ABEs provided herein exhibit reduced off-target editing effects while retaining high on-target editing efficiencies. These ABEs exhibit reduced off-target DNA editing effects and reduced off-target editing effects in cellular mRNA. In addition, methods for targeted nucleic acid editing are provided. Further provided are pharmaceutical compositions comprising the ABEs. Also provided are vectors and kits useful for the generation and delivery of the ABEs, including vector systems for engineering the ABEs through directed evolution. Cells containing such vectors and ABEs are also provided. Further provided are methods of treatment comprising administering the ABEs.

The present invention belongs to the field of genetic engineering. Specifically, the present invention relates to a directed evolution method based on geminivirus. More specifically, the present invention relates to a directed evolution method for in vivo screening of a genetic element in a plant cell by using primary and secondary replicons of geminivirus.

Methods of microbial screening for identifying alcohol acyltransferases for ester biosynthesis and submodules for ester pathways to produce butyryl-coenzyme A derived esters are disclosed. The method includes the introduction preselected plasmids into a respective host strain to form engineered microbes, in situ fermentation thereof followed by a colorimetric assay for quantification of production of the target ester. In situ fermentation includes inoculating each well of a microplate that have a culture media for producing target esters with one of the engineered microbes, adding an overlay of a solvent to each, and incubating the same. The colorimetric assay includes transfer of a quantity of the overlay from each well to respective clean wells of a new microplate, treatment of each well to form an iron-hydroxamic acid complex aqueous phase, centrifugation of the microplate, and measurement of the absorbance at 520 nm and comparison to a standard curve for the target ester.