An Argonaute protein from eukaryotes and an application thereof are provided. An amino acid sequence of the Argonaute protein is shown in SEQ ID NO: 1 or has at least 50% sequence identity with the sequence shown in SEQ ID NO: 1. The specific cleavage activity of the eukaryotic Argonaute protein on DNA is first proved, and an experimental proof for the study of interaction between the eukaryotic Argonaute protein and DNA is provided. In addition, polypeptides, nucleic acids, expression vectors, compositions, kits, and methods used therein can carry out site-specific operation on intracellular and extracellular genetic materials and can be effectively applied in many fields of biotechnology, providing a new tool for gene editing, modification, and molecular detection of Argonaute polypeptides based on eukaryotic sources.

A gene editing system of Escherichia coli includes an Escherichia coli, a helper plasmid and a donor plasmid. The helper plasmid successively includes a transposase complex expression cassette, a Cas12k expression cassette, a first sgRNA cassette, a first antibiotic resistance gene and a first replication origin. The donor plasmid successively includes a left end sequence of a ShCAST transposon, an exogenous gene expression cassette, a right end sequence of the ShCAST transposon, a second sgRNA cassette, a second antibiotic resistance gene and a second replication origin.

Compositions and use of the compositions in methods of detecting SARS-CoV-2 in a sample is disclosed, using RT-LAMP coupled with CRISPR-Cas12, referred to herein as iSCAN (in vitro Scanning of COVID-19-Associated Nucleic acids) is disclosed. iSCAN provides a rapid, specific, accurate, sensitive detection of SARS-CoV-2 in a sample. The iSCAN is 1) rapid, as the RT-LAMP and CRISPR-Cas12/Cas 13 reaction takes less than 1 h; 2) specific, because detection depends on the identification and subsequent cleavage of SARS-CoV-2 genomic sequences by the Cas12 or 13 enzyme; 3) field-deployable, as only simple equipment is required; and 4) easy to use, as the colorimetric reaction coupled to lateral flow immunochromatography makes the assay results easy to assess. The methods include amplifying SARS-CoV-2 in a sample using RT-LAMP and using the RT-LAMP product as a substrate in a CRISPR-Cas12/13 reaction, incorporated with a means of detecting the presence of the SARS-CoV-2 RT-LAMP product.

The present disclosure relates to genetically modified non-human animals that express a human or chimeric (e.g., humanized) LAG3, and methods of use thereof.

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.

Described herein are expression vectors encoding the Wolbachia protein WalE1. Also described are insects transformed with an expression vector of the present disclosure, and progeny thereof. Also described are methods for improving Wolbachia replication and transmission in its insect host by overexpressing WalE1 in the insect host. Improved Wolbachia replication and transmission provides for insect population control and pathogen resistance in insects, which can reduce disease transmission.

A Streptococcus canis Cas9 (ScCas9) ortholog and its engineered variants, possessing novel PAM specificity, is an addition to the family of CRISPR-Cas9 systems. ScCas9 endonuclease is used in complex with guide RNA, consisting of identical non-target-specific sequence to that of the guide RNA SpCas9, for specific recognition and activity on a DNA target immediately upstream of either an “NNGT” or “NNNGT” PAM sequence. A novel DNA-interacting loop domain within ScCas9, and other Cas9 orthologs, such as those from Streptococcus gordonii and Streptococcus angionosis facilitates a divergent PAM sequence from the “NGG” PAM of SpCas9.

The present invention relates a vector system and a vector system for use in a method of treating a disease, each comprising a first vector and a second vector. The present invention further relates to the first vector, the second vector and a combination of the first vector and the second vector. In addition, the present invention relates to a pharmaceutical composition comprising the vector system of the invention or the combination of the invention.

A nucleic acid construct is disclosed which is removable after transformation. Methods of using same are disclosed as well.

Some aspects of this disclosure provide strategies, systems, reagents, methods, and kits that are useful for engineering Cas9 and Cas9 variants that have increased activity on target sequences that do not contain the canonical PAM sequence. In some embodiments, fusion proteins comprising such Cas9 variants and nucleic acid editing domains, e.g., deaminase domains, are also provided.