Disclosed are a phosphinothricin dehydrogenase mutant, a recombinant bacterium and a one-pot multi-enzyme synchronous directed evolution method. The phosphinothricin dehydrogenase mutant, with an amino acid sequence as shown in SEQ ID No.1, is obtained by mutating alanine at position 164 to glycine, arginine at position 205 to lysine, and threonine at position 332 to alanine in a phosphinothricin dehydrogenase derived from Pseudomonas fluorescens. The recombinant bacterium is obtained by introducing a gene encoding the phosphinothricin dehydrogenase mutant into a host cell. The host cell can also incorporate a gene encoding a glucose dehydrogenase or a gene encoding a formate dehydrogenase to undergo synchronous directed evolution to achieve double gene overexpression. The one-pot multi-enzyme synchronous directed evolution method of the present invention can screen recombinant bacteria with greatly improved activity. Compared with other catalysis processes such as the transaminase method, the method for preparing L-PPT of the present invention features relatively simple process, high conversion of raw materials of up to 100%, and high stereo selectivity.

Disclosed herein are compositions, methods, and systems for selecting a polynucleotide for activity as a neuronal-specific transcription factor. The system may include a polynucleotide encoding a reporter protein and a pan-neuronal marker, a Gas protein, and a library of guide RNAs (gRNAs) targeting putative transcription factors. Further provided are methods of screening for a neuronal-specific transcription factor.

Cell populations, compositions, and methods are provided relating to target priming of macrophage cells. The macrophages, once primed or activated with a microorganism, can be used to prevent or treat infection by the microorganism. Likewise, once primed or activated by a tumor cell or tumor antigen, the macrophage cell can be used to prevent or treat tumor of the same kind. The priming can be carried out in vitro or ex vivo. The macrophages can be isolated from the subject of disease prevention or treatment.

Methods and compositions related to a nucleic acid-guided nuclease cell targeting screen are provided. The invention relates to compositions and methods for identifying cell targeting proteins that, when associated with a nucleic acid-guided nuclease (such as Cas9), enables at least the nucleic acid-guided nuclease to be targeted to the surface of a target cell or internalized by a target cell, i.e., a cell targeted by the cell targeting agent.

A method for identifying and characterizing biological parts based on omics datasets and a dual-fluorescent reporter gene system, and a biological part library constructed thereon are provided, relating to a technical filed of biology. The method includes steps of: identifying the biological parts using the omics datasets; constructing a single-fluorescent reporter gene system using a shuttle vector pEZ15Asp as a skeleton for screening and determining fluorescent reporter genes; obtaining a dual-fluorescent reporter gene system skeleton; constructing recombinant plasmids, and finally transforming into competent cells for quantitative analysis of fluorescence intensities. The present invention is convenient and quick, and can screen and identify different biological parts such as RBS, UTRs, promoters, and terminators of different intensities in batch quantitatively in a relatively short time. Moreover, the present invention can quickly expand the biological part library of Z. mobilis, so as to be applied in metabolic engineering of different demands.

A method for identifying and characterizing biological parts based on omics datasets and a dual-fluorescent reporter gene system, and a biological part library constructed thereon are provided, relating to a technical filed of biology. The method includes steps of: identifying the biological parts using the omics datasets; constructing a single-fluorescent reporter gene system using a shuttle vector pEZ15Asp as a skeleton for screening and determining fluorescent reporter genes; obtaining a dual-fluorescent reporter gene system skeleton; constructing recombinant plasmids, and finally transforming into competent cells for quantitative analysis of fluorescence intensities. The present invention is convenient and quick, and can screen and identify different biological parts such as RBS, UTRs, promoters, and terminators of different intensities in batch quantitatively in a relatively short time. Moreover, the present invention can quickly expand the biological part library of Z. mobilis, so as to be applied in metabolic engineering of different demands.

The present invention generally relates to bacterial polypeptide display systems, libraries using these bacterial display systems, and methods of making and using these systems, including methods for improved display of polypeptides on the extracellular surface of bacteria using circularly permuted transmembrane bacterial polypeptides that have been modified to increase resistance to protease degradation and to enhance polypeptide display characteristics.

Provided is method for producing a phage library displaying antibodies or antibody fragments, comprising providing a first polynucleotide containing LC, a second polynucleotide containing connexon, and a third polynucleotide containing HC, respectively introducing the first, second and third polynucleotides to first, second and third bacteria to obtain a light chain component bacterial library, a connexon component bacterial library, and a heavy chain component bacterial library, obtaining a light chain component plasmid, a connexon component plasmid, and a heavy chain component plasmid from the libraries, obtaining the released LC, released connexon and released HC from the plasmids, connecting the released display vector segments to form a connection product for display, introducing a third bacterium to obtain a display bacterial library, and using the display bacterial library to prepare the phage library for displaying the antibodies or antibody fragments. Also provided is a phage library produced according to the method.

The present application provides methods of enhancing T cell function (e.g., expansion, persistence and/or effector functions), particularly by genetic modification of the Regnase-1, Batf, and additional genes (alone or in combination). The application also provides modified T cells manufactured using the methods provided by this invention and related pharmaceutical compositions. The application further provides methods of using the modified T cells for treating a disease (e.g., a cancer or an infectious disease).

The present invention provides for a method for screening for gene function for a bacteriophage, the method comprising: (1) (a) providing one or more host organism, such as a species or strain, libraries, (b) providing randomly barcoded transposon sequencing (such as RB-TnSeq), and (c) screening for loss-of-function (LOF) mutant phenotypes; or (2) (a) providing one or more DNA barcoded overexpression strain libraries (such as Dub-seq) using DNA of the host organism and/or phage, and (b) screening for gain-of-function (GOF).