187 results about "Intron" patented technology

The invention provides an hpRNA interference vector for laver and application of the hpRNA interference vector, and belongs to the technical field of genetic engineering and algae biology. According to the hpRNA interference vector, a binary vector pBI121 is used as a skeleton vector, a laver endogenous Actin promoter and a multiple cloning site segment are inserted, the multiple cloning sites allow target gene segments to be cloned in a forward and reverse repetition mode, the target gene segments are spaced by introns, and therefore an efficient hpRNA structure is formed after transcription, and the efficient hpRNA interference vector is obtained. The gene can be directly used for silencing important genes related to growth and development, metabolic regulation, stress response and the like in the laver. The invention further provides a construction method of the hpRNA interference vector and application of the hpRNA interference vector in silencing laver genes and provides a method for silencing laver gene expression, the gene silencing effect is remarkable, experimental results can be repeated, and a powerful tool is provided for molecular breeding and genetic improvement of laver.

The invention relates to a high-throughput cis-acting element screening carrier and a screening method. Specifically, the invention provides a plasmid vector system containing bar codes, each bar code in the system is in one-to-one correspondence with a candidate cis-acting element, and the activation multiple of the candidate cis-acting element can be obtained by measuring the abundance of the bar codes; in order to eliminate the influence of a bar code on the vector on a detection result, an exogenous intron which can be cut off during transcription is inserted into a coding region in the vector and is used for distinguishing vector DNA and RNA obtained by transcription. The screening method of the biological cis-acting element has high efficiency, wide applicability and high throughput, and has outstanding application value in biological research and breeding.

The invention discloses a molecular marker combination related to a cockscomb height character and a detection reagent and application thereof, and belongs to the technical field of molecular genetics. According to the molecular marker combination and the detection reagent thereof, ten SNP loci remarkably associated with the chicken first-laying crown height and the 130-day-old crown height are identified in the intron of the CTCF gene for the first time, based on the SNP loci, the molecular marker combination related to the cockscomb height character and the detection reagent thereof are designed, and the molecular marker combination and the detection reagent thereof can be used for early screening of laying hens with high cockscomb height and can be used for early screening of laying hens with high cockscomb height. The method is of great significance to breeding and breeding of precocious high-quality laying hens.

The invention discloses an Indel marker related to a corn kernel protein content character and application of the Indel marker, and belongs to the technical field of plant molecular genetics and crop molecular design breeding. The Indel marker is located on a gdh2 gene of a chromosome 10 of a corn genome, and insertion or deletion of a nucleotide sequence as shown in SEQ ID NO.5 occurs in an intron region between a second exon and a third exon. By applying the molecular marker-assisted selection technology, high-protein new germplasms S3317 (the crude protein content is 17.0%) and JS231148 (the crude protein content is 15.6%) have been successfully cultivated, the increase amplitudes of the high-protein new germplasms and JS231148 are both more than 15% compared with those of the conventional control, and the lysine content reaches 0.41% and reaches the national high-quality protein corn standard.

The present invention relates to chimeric promoters without CpG motifs. The chimeric promoter comprises a CMV enhancer, an IFN-gamma gene core promoter and an intron sequence, and a CG motif in the CMV enhancer, the IFN-gamma gene core promoter and the intron sequence is subjected to substitution mutation. The promoter can improve the expression efficiency of exogenous genes in activated immune cells.

Provided is a method for producing a heterodimeric protein such as a bispecific antibody with which a heterodimeric protein composed of only domains having natural amino acid sequences can also be produced. A production method of the present invention is a method for producing a heterodimeric protein. The production method includes a production step of producing a heterodimeric protein by reacting a dimeric protein having a reaction tag with a modification protein for modifying the dimeric protein. The dimeric protein having a reaction tag includes a first monomeric protein and a second monomeric protein. The first monomeric protein includes a first reaction tag and a first dimer formation domain capable of forming a dimer in this order. The first reaction tag includes a binding tag and a first C intein. The second monomeric protein includes a second reaction tag and a second dimer formation domain capable of forming a dimer together with the first dimer formation domain in this order. The second reaction tag includes a binding partner capable of binding to the binding tag, and a second C intein. The modification protein includes a first modification protein and a second modification protein. The first modification protein includes a first N intein capable of reacting with the first C intein, and a first addition component to be added to the first monomeric protein. The second modification protein includes a second N intein capable of reacting with the second C intein, and a second addition component to be added to the second monomeric protein. The first addition component and the second addition component are different addition components. The first monomeric protein and the second monomeric protein form a dimer. In the production step, the first N intein of the first modification protein reacts with the first C intein of the first monomeric protein, and then the first addition component of the first modification protein is linked to the first monomeric protein, and the second N intein of the second modification protein reacts with the second C intein of the second monomeric protein, and then the second addition component of the second modification protein is linked to the second monomeric protein.

The invention discloses a CHRM3 gene InDel molecular marker associated with growth and reproduction traits of a large white pig and application of the CHRM3 gene InDel molecular marker, a 62bp deletion variation is identified in a second intron of the CHRM3 gene, and a breakpoint is positioned in a second intron 521426525142713bp (a pig chromosome 14 reference sequence NC010456.5) of ENSSSCT00000082112.1. The application for detecting / identifying the growth and reproduction traits of the large white pigs is developed on the basis of the molecular marker, and the detection method is simple, rapid and low in cost and can be used for accurately screening individuals with better traits such as backfat thickness, primary birth litter weight, total multiparous litter size and gestation period of multiparous sows in the early stage, so that genetic improvement is carried out on the growth and reproduction traits of the pigs, and the economic benefit is increased. Multiple characters such as growth and reproduction can be selected at the same time, the breeding period of excellent boars can be shortened, and the breeding efficiency and progress are accelerated.

The invention provides a single-chain and double-protein-expressed circular RNA construct and a preparation method and application thereof, and the single-chain and double-protein-expressed circular RNA construct comprises 5'and 3 'introns used for realizing RNA self-splicing and cyclization as a first kind of introns; the 5'and 3 'homologous arms are used as pairing sites in RNA molecules; the exon 1 and the exon 2, which are connected after being spliced, can be spliced and connected together under the action of the first type of introns to form a complete and continuous open reading frame; an interval sequence for providing spatial and adjustment sequence structures; the IRES is used for starting translation; a glycine-serine linker as a flexible linker; a green fluorescent protein for gene expression monitoring; a luciferase for producing light by a catalytic chemical reaction. All the advantages of the double-IRES system for generating natural complete protein are reserved, and the core defect of low efficiency can be thoroughly overcome.

Provided are a high-throughput transcriptome sequencing-based IKZF1 gene exon deletion recognition system and method. The exon deletion recognition system is obtained by inputting the number of IKZF1 exon junctions, differential genes and expression quantities thereof, and IKZF1 exon and intron mutation frequency information of samples as features into a constructed random forest model and performing prediction, and can be used for accurately predicting and recognizing any exon deletion with high accuracy and high specificity, providing guidance for patient prognosis and treatment.

The invention relates to an intron variant capable of accurately splicing and enhancing gene transcription and application of the intron variant, and belongs to the technical field of biological breeding, the intron variant is a variant of a Cat1mu intron, the Cat1mu intron has a nucleotide sequence as shown in SEQ ID NO.2, and the nucleotide sequence of the intron variant is formed by mutating base AG at the 182th to 183th sites of the sequence as shown in SEQ ID NO.2 into tG, Ac, At, cG, Aa, ta, tt, tc, cc, ct or ca. According to the present invention, the sequence analysis and the saturation mutation are performed on the Cat1mu, the screening is performed to obtain the Cat1mu series variant Cat1mu2-12, the accurate splicing effect of the Cat1mu2-12 during the fusion gene expression process is verified, and the Cat1mu6, the Cat1mu7, the Cat1mu8 and the Cat1mu10 can further enhance the fusion gene expression effect.

The invention relates to a full-length sequence amplification primer, an amplification method and a three-generation sequencing method of an HLA-I antigen gene, and belongs to the field of gene detection. The HLA-I antigen gene is subjected to PCR (Polymerase Chain Reaction) amplification by using 8 pairs of characteristic amplification primers. And each gene can be subjected to high-yield specific amplification on a full-length sequence containing complete intron and exon regions only through one round of amplification and enrichment, and 7.8 Kb long-fragment genes can be captured once at most. The experimental deviation caused by simultaneous amplification of a single gene by using a plurality of pairs of segmented primers and the indirect error caused by gene splicing are effectively avoided. The method comprises the following steps: carrying out quality inspection and purification on amplicon PCR (Polymerase Chain Reaction) products, mixing samples, constructing an HLA-I type antigen gene library, carrying out accurate and complete sequencing on the full-length sequence of the HLA-I type antigen gene by adopting a three-generation PacBio Sequel II platform, and ensuring that the Hifi reads base accuracy can reach 99% or above. The reading of HLA-I type antigen gene full-length sequence variation information and the haplotype analysis at a high resolution level can be realized.

The invention discloses a long-fragment PCR (Polymerase Chain Reaction) amplification reaction solution and application thereof. The PCR amplification reaction solution is composed of a 2XPCR buffer solution, dNTP, LA Taq enzyme, betaine and distilled water. In the PCR amplification reaction liquid, the volumes of the 2XPCR buffer solution, the dNTP, the LA Taq enzyme, the betaine and the distilled water in the total volume of 25l are respectively 12.5 l, 5.0 l, 0.25 l, 4.0 l and 0.45 l. According to the present invention, the PCR amplification reaction liquid can effectively amplify the long fragment PCR product, the product amplification with the longest detection reaching 12 kb can be used for the detection of the hemophilia A intron 22 inversion, the effective amplification reagent is provided for the long fragment PCR amplification, and the problem of difficult long fragment amplification is solved.

A method and compound for regulating SCN1A gene mRNA and protein expression. Provided is a use of a guide agRNA in preparation of a drug for altering expression of a target protein in a cell of a subject. The cell has a precursor mRNA encoding the target protein, wherein the precursor mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or a pseudo 3' splice site of the intron; the guide agRNA is capable of binding to the precursor mRNA to form a structure capable of recruiting ADAR in the cell, thereby forming a complex containing an ADAR protein; and the target protein is a Nav1.1 protein. By regulating alternative splicing of a target gene pre-mRNA, the functional RNA of the target gene and the function or expression of the target protein are affected, thereby achieving the objective of disease treatment.

Abstract Methods for editing, e.g., introducing a heterologous transgene, within the human albumin gene (e.g., at intron 1) are provided. Abstract

Lung cancer remains the leading cause of cancer-related deaths in the world. Despite the fact that introduction of immune checkpoint inhibitors (ICIs) led to a major advancement in lung cancer treatment, disease prognosis continues to remain low and a significant proportion of patients do not respond to such therapies. Cancer vaccines could potentially provide a complementary approach to boost antitumor immunity and act synergistically with ICIs. Novel tumor antigens shared by a large proportion of lung tumor cells are described herein. Several of the tumor antigens described herein derive from aberrantly expressed unmutated genomic sequences, such as intronic and intergenic sequences, which are not expressed in normal tissues. Nucleic acids, compositions, cells and vaccines derived from these tumor antigens are described. The use of the tumor antigens, nucleic acids, compositions, cells and vaccines for the treatment of lung cancer is also described.

The present invention relates to methods and devices for identification and quantification, including low abundance nucleotide base mutations, insertions, deletions, translocations, splice variants, miRNA variants, alternative transcripts, alternative start sites, alternative coding sequences, alternative non-coding sequences, alternative splicing, exon insertions, exon deletions, intron insertions, or other rearrangements and / or methylated nucleotide bases at the genomic level.

The invention relates to the technical field of animal breeding, in particular to a method for breeding black sheep. The method comprises the following steps: introducing an expression cassette into a cell to obtain a transgenic cell; and carrying out somatic cell nuclear transplantation cloning by adopting the transgenic cells to obtain the black sheep. The expression cassette comprises an intron of PK5 and beta-globin, and an EDN3 gene, the PK5 comprises a nucleotide sequence as shown in SEQ ID NO. 2; and the EDN3 gene comprises a nucleotide sequence as shown in SEQ ID NO. 4. A specific expression cassette and a carrier are researched, the black sheep of which the skin, hair, tongue, oral mucosa, eye white mucosa, epididymis and ears all become black can be prepared through somatic cell cloning on the basis of cells edited by the carrier, and important application value and economic value are achieved.

The present application relates to methods of forming circularized RNA (circRNA) using intact Group I introns provided in cis or fragments of Group I introns provided in trans. The present application also relates to methods of forming circRNA using intact Group II introns provided in cis or fragments of Group II introns provided in trans.

Recombinant polynucleotides and vectors containing an engineered (artificial) exon-intron-exon gene structure in a transgene are provided, which undergoes splicing when it is expressed in a target cell.

This invention belongs to the field of gene technology for disease control. This invention provides a method for preparing biocontrol engineered bacteria using *Trichoderma harzianum* as a dsRNA vector, comprising the following steps: determining the target gene based on the pathogenic fungus; using the target gene as an arm sequence, and centering on the VdTublin intron sequence of the *Verticillium dahliae* endogenous gene, connecting two identical arm sequences to both ends of the intron in a forward and reverse direction, respectively, to construct the target sequence; and transferring the target sequence into *Trichoderma harzianum* using ATMT to obtain engineered *Trichoderma harzianum* bacteria. The engineered strain obtained by this invention has a good inhibitory effect on the growth and pathogenicity of the corresponding fungus.

The invention discloses an SNP (Single Nucleotide Polymorphism) marker related to the body size and temperature adaptability of worker bees of apis cerana and application of the SNP marker. According to the invention, 25 key SNP sites and 10 associated genes for regulating and controlling the body type and temperature adaptation of the apis cerana are screened by a selection signal analysis method. Research finds that in the second intron region (6452066th site of the chromosome 7 of the bees) of the Foxo gene, the mutation type is T / C, and the mutation type is closely related to the body size and temperature adaptability of the Chinese bees. Through verification, in Hainan apis cerana and Aba apis cerana with extremely different body sizes and living environment temperatures, mutation position genotypes are different, TT is a large-body cold-resistant dominant genotype, and CC / TC is a small-body heat-resistant dominant genotype; a gene expression quantity detection result shows that the expression quantity of the Foxo gene is in negative correlation with a large body type and cold resistance and is in positive correlation with a small body type and heat resistance. The method can be used for genetic improvement of bee body shape characters and has breeding application value.

The application provides a genetic marker associated with chicken intestinal length in a KCNIP4 gene and an application thereof, and belongs to the fields of animal genetics and breeding and biotechnology.The genetic marker comprises IL_tag1 or IL_tag2; the Ensembl number of the IL_tag1 is rs316532738, corresponds to the sequence of a positive strand of a chromosome No.4 of a chicken reference genome bGalGal1.mat.broiler.GRCg7b published by NCBI, is located in the 1st intron of a gene KCNIP4, and the base at the position is T or C; the Ensembl number of the IL_tag2 is rs316953671.The genetic marker is helpful to genetically improve the intestinal length of a laying hen, is applied to the genetic breeding of a chicken, and is favorable to improving intestinal traits and obtaining a laying hen variety with better nutrient absorption.

A method for regulating the mRNA and protein expression of an SCN1A gene, and a compound. Provided is a use of an antisense oligonucleotide guide RNA in the preparation of a drug for altering the expression of a target protein in a cell of a subject. The cell has a pre-mRNA encoding the target protein, wherein the pre-mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or pseudo 3' splice site of the intron; the antisense oligonucleotide guide RNA can bind to the pre-mRNA to form a structure capable of recruiting ADAR in a cell, thereby forming a complex containing an ADAR protein; and the target protein is an Nav1.1 protein. That is, the function or expression of the functional RNA and target protein of a target gene is affected by regulating the alternative splicing of the pre-mRNA of the target gene, thereby achieving the objective of treating diseases.

The invention provides a molecular marker, a detection reagent and a method for evaluating growth traits and / or egg laying interval traits of meat pigeons. The SNP locus and the haplotype molecular marker which are remarkably related to the body size character, the body weight character and / or the egg laying interval character of the meat pigeon are screened from the first intron of the ASIP gene of the meat pigeon, and can be applied to early auxiliary selection of meat pigeon breeding. The invention further provides a detection reagent and a detection method for detecting the SNP loci and the haplotype molecular markers, the body size character, the body weight character and / or the egg laying interval character of the meat pigeons can be quickly, effectively and accurately predicted, so that the production cost is saved, the breeding process is accelerated, and the detection reagent and the detection method have high economic application value and scientific research value.

The invention relates to the field of molecular biology, and further relates to a specific marker, a primer pair, a detection method, a kit and application of macal2 gene insertion variation of oplegnathus fasciatus. The specific marker is a DNA (Deoxyribose Nucleic Acid) fragment in which the specific marker is not inserted and inserted in the intron region of the macal2 gene of the striped sea bream, and the nucleotide sequences of the DNA fragment are as shown in SEQ ID NO: 1 and SEQ ID NO: 2. According to the invention, through a pair of specific primers, a 118bp band and a 391bp band are amplified in an individual with DNA insertion variation, and only the 118bp band is amplified in an individual without insertion variation. According to the detection method, the identification time is remarkably shortened, the detection efficiency is improved, and the method has important application value in sex identification, high-male fry preparation and family breeding of oplegnathus fasciatus.

The present disclosure provides DNA molecules, such as transfer plasmids, useful for manufacturing packaged retroviral vectors encoding a therapeutic protein such as chimeric antigen receptors (CARs) and T cell receptors (TCRs). The encoded retroviral transcript in the DNA molecule includes a transgene, an elongation factor 1-alpha 1 (EFla) promoter, without the EFla intron, operably linked to the transgene and a Rev response element (RRE).

The invention discloses a method for preparing a CD163 gene editing pig based on a CRISPR (clustered regularly interspaced short palindromic repeats) / Gs12-7MAX system. The method specifically comprises two editing strategies: one is to edit a sixth intron and a seventh intron of a CD163 gene to realize accurate deletion of a seventh exon so as to obtain a porcine fibroblast monoclonal cell with homozygous deletion of the seventh exon; and 2, editing aiming at a seventh exon of the CD163 gene, and inducing the exon to generate frame-shift mutation to obtain a gene editing pig. The technical system established by the invention has the advantages of high editing efficiency, strong accuracy and simplified operation process, provides a new effective strategy for prevention and control of PRRSV (porcine reproductive and respiratory syndrome), and has important popularization and application values in creation of new materials for pig disease-resistant breeding.

The present application relates to MINI-REP gene for evaluating USH2A exon 13 knockout efficiency and its expression vector and application, and relates to the technical field of genetic engineering. The MINI-REP gene comprises a reporter gene and a small gene, wherein the reporter gene is a gene for performing a reporting function, and the small gene comprises genes of USH2A intron 12, exon 13 and intron 13 connected in sequence; the small gene is inserted into the reporter gene, thereby dividing the reporter gene into split reporter genes, which do not perform the reporting function when expressed alone, but perform the reporting function when expressed in series. The MINI-REP gene can be applied to efficiency evaluation of all related technologies for inducing USH2A exon 13 knockout in the prior art, and can be used for research and development of new technologies, effect detection, efficient screening, rapid, intuitive, efficient and high-sensitivity qualitative and quantitative analysis of USH2A exon 13 knockout efficiency.

The invention discloses a sequence optimization method for improving the stability of circular RNA, application and a product, and belongs to the technical field of biotechnology and molecular biology. In order to solve the problem that the existing circular RNA is easy to be specifically degraded by RNAseK and lysosome after entering a cell, synonymous mutation optimization is carried out on an open reading frame of a target gene of the circular RNA by identifying nucleic acid motifs (a first type of nucleic acid motifs and a second type of nucleic acid motifs) identified by two degradation mechanisms. The optimized circular RNA is prepared by means of in-vitro cyclization of an anabaena I-type intron self-splicing mediator, after cells are transfected, the intracellular RNA expression level of the circular RNA is remarkably improved compared with that of a wild type, and the target protein expression efficiency is also remarkably improved. The intracellular stability and expression efficiency of the circular RNA are remarkably enhanced, and the method can be widely applied to the scenes of gene expression regulation, protein synthesis, gene therapy, vaccine development and the like.

The invention relates to the technical field of animal breeding, in particular to an expression vector based on a skin specific promoter and application of the expression vector in preparation of black skin pigs. The expression vector comprises an expression cassette, and the expression cassette comprises a 5'end regulation region of a pig source K5 gene, an intron of a beta-globin gene, a target gene and a 3 'end regulation region of a K14 gene. An expression vector for specifically expressing a target gene in animal skin is provided by adopting a regulatory region of a porcine K5 gene and an intron of a beta-globin gene, and the target gene (such as a melanin synthesis regulatory gene) is integrated by adopting the expression vector and can be efficiently expressed in a porcine somatic cell so as to prepare a pig with specific characters; the method has an important application value in the pig raising industry.