36 results about "Insertion site" patented technology

Compositions and methods related to transgenic glyphosate tolerant Brassica plants are provided. Specifically, the present invention provides Brassica plants having a DP-073496-4 event which imparts tolerance to glyphosate. The Brassica plant harboring the DP-073496-4 event at the recited chromosomal location comprises genomic / transgene junctions within SEQ ID NO: 2 or with genomic / transgene junctions as set forth in SEQ ID NO: 12 and / or 13. The characterization of the genomic insertion site of the event provides for an enhanced breeding efficiency and enables the use of molecular markers to track the transgene insert in the breeding populations and progeny thereof. Various methods and compositions for the identification, detection, and use of the event are provided.

This invention relates to the field of bioengineering technology, specifically to a recombinant adenovirus displaying a large antigen fragment on its capsid surface, its construction method, and its applications. The HVR1 and HVR5 regions of the hexon hypervariable region of this virus are replaced by gene fragments containing exogenous genes; each exogenous gene fragment has a flexible linker attached to both ends, and the gene sequence of the flexible linker is shown in SEQ ID No. 1. By precisely locating the naturally variable HVR1 and HVR5 loop regions exposed on the surface of the hexon protein as insertion sites, and supplementing them with flexible linkers, the core problem of exogenous sequence insertion disrupting the stability of the hexon structure is successfully solved. This allows for stable packaging to produce viral particles with high titers and infectivity, enabling the development of a new generation of highly efficient and safe capsid-displaying adenovirus vaccines.

The invention belongs to the field of molecular biology and genetic engineering, and discloses a chemiluminescent reporter plasmid pBBR402 and application thereof in detection of the transcription level of a target gene of acetobacter pasteurianus. The plasmid comprises a replication element (oriV and rep) from a pBBR1MCS-2 plasmid, a kanamycin resistance gene from a pMS402 plasmid, a promoter insertion site (BamHI / AvaI / XhoI), a luxCDABE reporter gene cluster and a T1 transcription terminator, and the kanamycin resistance gene, the promoter insertion site (BamHI / AvaI / XhoI), the luxCDABE reporter gene cluster and the T1 transcription terminator are obtained from the pBBR1MCS-2 plasmid. The plasmid can be inserted into a to-be-detected gene promoter of acetobacter pasteurianus to drive luxCDABE expression. During application, the recombinant plasmid is transferred into acetobacter pasteurianus, the OD value and the LUM value of the living bacteria liquid are directly detected, and the transcription level is quantified by the LUM / OD value. The method is easy to operate, high in sensitivity and suitable for gene regulation and control research of acetobacter pasteurianus.

A modified vector of adenovirus-associated virus serotype 8 (AAV-8) for gene targeting and expression is provided, wherein the modified vector includes serotype coat amino acid sequence, insertion site and insertion amino acid sequence. A 10-amino acid sequence set forth in SEQ ID NO.3 is inserted between amino acids at positions 590 and 591 that are set forth in SEQ ID NO.2 of the AAV-8 serotype coat protein: LARGDSTKSA, wherein amino acids at positions 1, 2 and 10 are protective amino acids, and amino acids at positions 3 to 9 are screened amino acid sequences. In addition, the present invention also discloses construction method and application of the modified vector are also provided.

This disclosure relates to bicistronic polypeptide constructs for use in allogeneic gene therapy, such as CAR-T cell therapy. A bicistronic construct comprises a first polynucleotide encoding a therapeutic molecule (e.g., CAR-T or antibody) and a second polynucleotide encoding an immune surveillance masking molecule (ISMM). The ISMM includes a non-functional version of a protein knockout, such as a human leukocyte antigen E genetically fused to a fragment, by insertion of a bicistronic construct, such as beta-2 microglobulin or B2M. Also provided are vectors comprising bicistronic constructs, cells (e.g., CAR-T cells), and methods of use. Kits and manufacturing articles are also provided. This disclosure also provides four novel insertion sites that may be used to insert expression constructs into the B2M gene.

The invention relates to the technical field of genetic engineering, in particular to identification of a new insertion site of a self-assembled protein cage nanoparticle antigen epitope and application of the new insertion site in vaccine preparation. It is found that a foot-and-mouth disease virus antigen epitope is inserted between 62nd-63rd amino acids of an Encapsulin protein fragment after amino acid sequence modification, and / or the 62nd-63rd amino acids are replaced with the foot-and-mouth disease virus antigen epitope, efficient and soluble expression of a target antigen in escherichia coli can be achieved, and protein cage nano antigen particles are successfully self-assembled; the protein nano antigen particle can induce a widely neutralized anti-foot-and-mouth disease virus antibody, improves the immune efficacy, and has the potential of becoming a foot-and-mouth disease vaccine.

The application discloses a rapid detection method of integration hot spots based on exogenous genetic material insertion sites, uniquely encodes chromosome-site information of all ISs to obtain a digital Y representing the chromosome and the integration position; sorts Ys corresponding to all ISs; for each IS in the insertion site data set, firstly, records the IS as a CIS node C; records other nodes Vi in the insertion site data set; if the distance between C and Vi is less than a threshold TH, records Vi added to the current CIS to become a node of the CIS; simultaneously, connects the edge of C and Vi; then, continues to return to step 2, continues the next IS, and sequentially obtains the CIS network; based on the connection between all ISs, the CIS network is searched to search for a CIS meeting the condition; the application optimizes the connection searching mode and the CIS searching range, and in the detection of several integration sites, the average analysis time of the software is greatly reduced, and the clinical practicability of the CIS detection algorithm is increased.

This invention belongs to the field of biotechnology, specifically relating to a *Pseudomonas putida* genome insertion site library and its construction method. This invention, for the first time, systematically screened and obtained 17 specific genome insertion sites suitable for *Pseudomonas putida*. The constructed insertion site library covers different expression intensities, filling the gap in existing technologies lacking efficient integration sites for *Pseudomonas putida*, and providing diverse options for personalized expression of target genes. The screened insertion sites are all located in the regions of opposing transcription genes, avoiding interference with the host's basal metabolic network. The recombinant strain exhibits stable growth performance without significant growth inhibition, making it suitable for large-scale industrial applications. The integration method, based on homologous recombination, achieves stable integration of the target gene without plasmid maintenance, avoiding expression instability caused by plasmid loss and copy number fluctuations. It also reduces the continuous use of antibiotics, lowering industrialization costs and environmental risks, and the construction method is simple and efficient.

The present disclosure relates to a method of detecting an integration site of a vector in a genome. According to the method of the present disclosure, it is possible to simply and quickly analyze a quantitative integration site of a viral vector with respect to a plurality of DNA motifs (sites) in a genome. Therefore, it can be useful for monitoring the safety and effectiveness of a gene therapy agent.

The invention discloses an eurytopic chlamydomonas reinhardtii expression vector based on a PSAD promoter and a 3 * Flag tag as well as a construction method and application of the eurytopic chlamydomonas reinhardtii expression vector. The nucleotide sequence of the eurytopic chlamydomonas reinhardtii expression vector is shown as SEQ ID No.1; the vector comprises a PSAD promoter, a target gene insertion site EcoR V, a 3 * Flag tag at the 3'terminal, an rbcS2 terminator, a TUB2 promoter, a hygromycin B screening gene and an rbcS2 terminator. The construction method comprises the following steps: constructing a skeleton of a pHyg-PSAD-3Flag vector; constructing a sequence fragment fused with a 3 * Flag tag; and connecting the sequence fragment fused with the 3 * Flag tag with a plasmid skeleton fragment through T4 DNA (deoxyribonucleic acid) enzyme so as to construct the pHyg-PSAD-3Flag vector. According to the invention, the expression plasmid of the chlamydomonas pHyg-PSAD-3Flag can be used for efficiently screening algae strains capable of stably expressing target protein.

This application provides a protein stability detection probe based on ascorbate peroxidase 2 and its application. The detection probe includes ascorbate peroxidase 2 displayed on the cell surface and an anchoring protein that anchors ascorbate peroxidase 2 to the cell. Ascorbate peroxidase 2 has an insertion site for the target protein, into which the target protein inserts. Ascorbate peroxidase 2 is divided into N-terminal and C-terminal ascorbate peroxidase 2, and the target protein is linked to both ends of the C-terminal ascorbate peroxidase 2 via flexible linkers. The intensity of the fluorescence signal generated by the cascade reaction catalyzed by the probe in this application has a good linear relationship with the stability of the inserted test protein mutant, enabling high-throughput identification of protein mutant stability and showing promising application prospects.

The invention relates to the technical field of genetic engineering, in particular to Encapsulin fusion protein for expressing foot and mouth disease virus epitopes and application of the Encapsulin fusion protein in preparation of subunit vaccines. The invention discovers that different foot-and-mouth disease virus antigen epitopes are inserted among the 62nd to 63rd positions, the 124 to 125th positions, the 138 to 139th positions and the 239th to 240th positions of amino acids of an Encapsulin protein fragment subjected to amino acid sequence modification, and / or the 62nd to 63rd positions, the 124 to 125th positions, the 138 to 139th positions and the 239th to 240th positions of amino acids are replaced by different foot-and-mouth disease virus antigen epitopes; efficient and soluble expression of a target antigen in escherichia coli can be realized, and protein cage nano antigen particles are successfully self-assembled; the protein nano antigen particle can induce a widely neutralized foot-and-mouth disease virus antibody, improves the immune efficacy, and has the potential of becoming a broad-spectrum foot-and-mouth disease vaccine.

A novel transgenic corn elite event designated MZIR098 is disclosed. The invention relates to DNA sequences of the recombinant constructs inserted into the corn genome and of genomic sequences flanking the insertion site that resulted in elite event MZIR098. The invention further relates to assays for detecting the presence of the DNA sequences of corn elite event MZIR098, to corn plants and corn seeds comprising the genotype thereof, and to methods for producing a corn plant by crossing a corn plant comprising the elite event MZIR098 genotype with itself or another corn variety.

Polypeptides are disclosed having an amino acid sequence at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence selected from SEQ ID NO: 1-28, not including any amino acid insertions at identified insertion sites, wherein the polypeptide binds to the F 1 domain of insulin receptor; or SEQ ID NO: 29-31, wherein the polypeptide binds to the LI domain of insulin receptor; fusion proteins thereof, and. methods for their use.

The invention discloses a herbicide-resistant alfalfa transformation event TWMS2-17 and a detection method thereof, an exogenous DNA fragment is inserted into an alfalfa genome, and herbicide-resistant plants TWMS2-17 are obtained through herbicide spraying treatment and single-copy plant screening. Through analysis and verification of a flanking sequence of a T-DNA insertion site, the position of an exogenous gene inserted into a genome is a No.3 chromosome, a 270bp genome sequence is replaced, and after asexual cuttage planting, excellent glyphosate and glufosinate resistance is shown, and normal growth is realized. The DNA molecule of the transformation event and the detection method have remarkable application value on breeding of herbicide-resistant alfalfa.

The invention discloses a high-throughput transgene insertion site detection method based on a next-generation sequencing technology. The method is based on the sequencing library obtained by the construction method disclosed by the invention, and the construction method of the sequencing library comprises the following steps: (1) extracting a transgenic positive sample whole genome; (2) fragmenting the genome DNA, and purifying to obtain a first mixture; (3) carrying out PCR (Polymerase Chain Reaction) amplification on the first mixture by adopting a specific primer with a UMI sequence; (4) taking the purified first round of amplification product as a template, mixing the template with the primers ad1-F and ad1-R, and performing second round of amplification; and (5) uniformly mixing a certain amount of the second round of amplification products, taking the sample mixed product as a template, mixing the sample mixed product with the primers ad2-F and ad2-R, and carrying out third round of PCR amplification. The method provided by the invention overcomes the defects of low flux, low efficiency and high cost of the existing transgenic insertion site detection method, and realizes deep sequencing and analysis of the flanking sequence of the insertion site in combination with a next-generation sequencing technology.

A method of transposon insertion sequencing (TIS) capable of resolving insertion sites in or around repeat elements, said method including the steps of; preparing a pool or library of mutant cells by

The invention provides brinjal event EE-6726, and plants, plant cells, seeds and plant parts comprising event EE-6726 which confers resistance to Lepidopteran insect damage. The invention also provides nucleic acid sequences specific for event EE-6726 and plants, plant cells, seeds and plant parts comprising nucleic acids specific for event EE-6726. This invention also provides methods for detection the presence of the event EE-6726 based on DNA sequence of the recombinant construct inserted into the brinjal genome that resulted in the EE-6726 event and / or the genomic sequences flanking the insertion site.

Disclosed are patch devices, methods, kits and systems for reducing the incidence of infections associated with the insertion site of a transdermal medical device (e.g., blood stream infections, infections of the skin opening through which the device enters the body, CLABSI, CRBSI) for example, the insertion site where a cannula or catheter enters a living body by generating an electromagnetic antiseptic signal in proximity of the insertion site.

The present invention relates to recombinant Marek's disease viruses comprising at least one recombinant nucleotide sequence encoding an antigen, operably linked to a promoter and an insulator, inserted into at least one insertion site, their production, compositions containing them, and their uses.

The compositions and methods disclosed relate to DNA compositions, plant cells, seeds, plant parts that relate to maize plants with increased grain yield trait. Also provided are assays for detecting the presence of the maize DP-202216-6 event based on the DNA sequence of the recombinant construct inserted into the maize genome and the DNA sequences flanking the insertion site. Kits and conditions useful in conducting the assays are provided.

An improved baculovirus expression system for enhanced heterologous protein expression in hosts selected from Bombyx mori larvae, pupae, primary cells, or established insect cell lines is disclosed. The expression system is derived from a wild-type strain of Bombyx mori nucleopolyhedrovirus (BmNPV) by deletion or inactivation of non-essential genes, of the BmNPV genome. The system includes novel transfer vectors designed for targeted gene modification and incorporation of a bacterial origin of replication and an insertion site for a gene of interest (GOI). This engineered bacmid maintains infectivity while offering enhanced genetic stability and significantly improved heterologous protein expression in insect cells, larvae, or pupae. Methods for constructing this bacmid via homologous recombination and using it for high-level protein production are also described.

Synthetic MVA-based vaccine compositions for preventing or treating a virus infection such as a coronavirus infection and methods of producing the vaccines. The vaccine compositions include (i) either a single DNA fragment that includes the entire genome of MVA, or two or more DNA fragments, each including a partial sequence of the genome of the MVA such that the two or more DNA fragments, when expressed in the host cell upon co-transfection, are assembled sequentially and comprise the full-length sequence of the MVA genome, and (ii) one or more DNA sequences encoding one or more human coronavirus antigens, subunits, or fragments thereof inserted in one or more insertion sites of the MVA. The antigens, subunits, or fragments thereof are expressed in the host cell upon transfection of the one or more DNA fragments.

The invention belongs to the technical field of gene engineering, and provides a transposon insertion Pro-TE closely linked with peach late flowering traits and application thereof.The transposon is obtained from peach tree PpTRX1 gene promoter region cloning, and the nucleotide sequence of the transposon is shown as SEQ ID NO.1; when the two peach varieties are used for treating the instantaneous transgenic tobacco plant, the transposon insertion type promoter Pro-TE increases the expression level of the gene LUC, which shows that the LUC fluorescence signal intensity of the transgenic plant is obviously improved; the discovery proves that Pro-TE plays a key role in regulating and controlling the flowering time of plants, and a new molecular target is provided for regulating and controlling the flowering period of peach trees; the transposon insertion site and the derived promoter sequence thereof can be used as a molecular marker co-segregated with late flowering characters, provide an important tool for peach tree molecular marker assisted breeding, and have important application values for cultivating anti-frost late flowering varieties and improving the stress resistance of the peach industry.

The present disclosure discloses a genome-wide insulator screening system. The system comprises an MAI-seq-experiment vector and an MAI-seq-control vector; core gene elements in the MAI-seq-experiment vector are arranged in following order: a weak promoter, a marker target gene, an insertion site for a sequence to be screened, an enhancer and a poly A site; and core gene elements in the MAI-seq-control vector are arranged in following order: a weak promoter, a marker target gene, an enhancer, an insertion site for the sequence to be screened and a poly A site. This genome-wide insulator screening system, primarily composed of these two vectors, exhibits high sensitivity and can be applied to the screening of genomic insulators in any species. Furthermore, it effectively eliminates the influence of silencers, ensuring high screening accuracy. This system provides important technical support for constructing insulator maps and understanding the characteristics and mechanisms of action of insulators.

The invention belongs to the technical field of marine organisms, and relates to an LAMP (loop-mediated isothermal amplification) primer probe combination and a kit for identifying a gracilaria lemaneiformis culture strain 981 and application of the LAMP primer probe combination and the kit, the nucleotide sequence of FIP of an inner primer is as shown in SEQ ID NO.1, the nucleotide sequence of BIP of the inner primer is as shown in SEQ ID NO.2, the nucleotide sequence of F3 of an outer primer is as shown in SEQ ID NO.3, and the nucleotide sequence of B3 of the outer primer is as shown in SEQ ID NO.4. The method is applied to large red alga strain identification for the first time, a four-primer LAMP system is developed on the basis that two sites are inserted into one InDel specific to the 981 strain, and during LAMP amplification, FIP primer modification biotin and BIP primer labeling FAM are combined with a streptavidin detection line of an LFD test strip, so that naked eye interpretation without electrophoresis is realized, the identification time is greatly shortened, and on-site rapid detection can be performed.

The invention discloses a transposon insertion site identification method based on high-throughput sequencing and application. A transposon with high copy and high sequence consistency is obtained through genome repetitive sequence analysis, restriction enzyme for library construction is determined through enzyme cutting site analysis, and a transposon terminal enriched library is constructed through the steps of genome DNA enzyme cutting, joint connection, PCR amplification and the like. Flanking sequence information of the transposon insertion site is obtained through high-throughput sequencing, the position of the insertion site on a genome is positioned through comparison with a reference genome, primers at the two ends of the insertion site are designed for identification of the insertion site, and therefore the high-throughput sequencing-based transposon insertion site identification method is established. According to the invention, the transposon terminal enriched library can be effectively constructed. Through high-throughput sequencing and bioinformatics analysis, high-throughput identification of transposon insertion sites can be realized.

This invention discloses a TaSMT-5B gene for enhancing wheat resistance to Fusarium head blight and its applications. The TaSMT-5B gene belongs to the selenocysteine ​​methyltransferase family, catalyzing the methylation of SeCys to MeSeCys. It is stably and highly expressed in the high-MeSeCys-rich and Fusarium head blight-resistant variety Zheng 9023, and a "GTCGCCG" insertion site exists in the promoter region of this gene in the Zheng 9023 variety. The TaSMT-5B gene was overexpressed using a constructed overexpression vector and introduced into the popular variety Fielder via Agrobacterium infection, resulting in five stable transgenic lines. After basal application of inorganic selenium, the severity of Fusarium head blight in the transgenic plants was alleviated. This invention has significant production implications for achieving selenium biofortification in wheat and enhancing its resistance to Fusarium head blight, providing a new gene resource for breeding stable-yielding and high-quality wheat varieties.

The present invention relates to recombinant Marek's disease viruses containing, inserted into at least one insertion site, at least one recombinant nucleotide sequence encoding an antigen operably linked to a promoter and an insulator, their manufacture, compositions comprising the same, and the uses thereof.

An improved baculovirus expression system for enhanced heterologous protein expression in hosts selected from Bombyx mori larvae, pupae, primary cells, or established insect cell lines is disclosed. The expression system is derived from a wild-type strain of Bombyx mori nucleopolyhedrovirus (BmNPV) by deletion or inactivation of non-essential genes, of the BmNPV genome. The system includes novel transfer vectors designed for targeted gene modification and incorporation of a bacterial origin of replication and an insertion site for a gene of interest (GOI). This engineered bacmid maintains infectivity while offering enhanced genetic stability and significantly improved heterologous protein expression in insect cells, larvae, or pupae. Methods for constructing this bacmid via homologous recombination and using it for high-level protein production are also described.