113 results about "Operon" patented technology

The invention relates to the technical field of biology, in particular to a molecular switch system with a signal cascade amplification function and application of the molecular switch system. The invention provides a molecular switch of a signal cascade method and a gene expression or regulation system. The molecular switch comprises an activator protein araC gene, a bidirectional pTet promoter, a repressor protein tetR gene, a pBad promoter and a tetO operon which are connected from upstream to downstream. When the molecular switch is used for expression or silencing of a target gene, lower background leakage, higher response rate, larger expression range and higher expression intensity are achieved, and expression is more homogeneous; and the inhibition efficiency is obviously improved.

The invention provides a lysine artificial operon and application thereof. The gene sequence of the artificial operon comprises a promoter, a lysC gene for coding aspartate kinase, an asd gene for coding aspartate semialdehyde dehydrogenase, a dapA gene for coding dihydropicolinic acid synthetase and a dapB gene for coding dihydropicolinic acid reductase, the ddh gene is used for coding diaminopimelate dehydrogenase; the lysA gene is used for coding diaminopimelate decarboxylase; except for the first lysC gene, sequences of RBS ribose binding sites of other genes are RBS ribose binding sites translated after gene transcription by intercepting 20-26nt DNA sequences at the upstream of a coding region ATG (adenosine triphosphate); the RBS ribose binding site is connected in front of the corresponding gene.

The invention belongs to the field of metabolic engineering, and discloses a genetically engineered bacterium of high-yield beta-alanine as well as a construction method and application of the genetically engineered bacterium. The genetically engineered bacterium takes escherichia coli W3110 as an original strain, and is obtained by performing the following gene editing on a genome of the escherichia coli W3110, the genetically engineered bacterium does not express proteins such as lactose operon repressor protein, pyruvate dehydrogenase, acetokinase and the like; protein such as aspartic acid-alpha-decarboxylase, phosphoenolpyruvate carboxylase, pyruvate carboxylase and the like is overexpressed; the phosphoenolpyruvate carboxykinase is replaced by phosphoenolpyruvate carboxykinase from Mannheim succinic acid producing bacteria. The genetically engineered bacterium BA-17 for high yield of beta-alanine, which is constructed by the invention, is fermented in a 5L fermentation tank for 60 hours, the yield of beta-alanine reaches 126.7 g / L, the conversion rate reaches 60.3%, and the production intensity reaches 2.11 g / L / h.

The invention provides a valine production strain and a construction method and application thereof, a designed acetohydroxy acid synthase mutant is that the 88th basic group of an ilvB gene is changed from a to c, the 382nd basic group is changed from a to g, the 413th basic group is changed from c to t, the gene sequence of a designed artificial operon comprises a promoter, an ilvB (A138V) gene or ilvB (Q30K, S128G, A138V) gene of coding mutated acetohydroxy acid synthase, and an ilvN (G20D, I21D, I21D, I21D, I21D, I21D, I21D, I21D) gene. I22F) gene, a pyk gene for coding pyruvate kinase, and a terminator; by designing a specific acetohydroxyacid synthase mutant and related biological materials and artificial operon, the strain constructed by directional modification of the strain by using a pK18mobsacB system gene editing technology based on allele exchange has the advantages of good genetic stability, high fermentation yield and the like, and valine can be stably produced.

The invention provides a valine production strain as well as a construction method and application thereof, the strain is constructed by utilizing a CRIPSR / Cas9 system gene editing technology based on allele exchange, an artificial operon is designed and constructed, the artificial operon is constructed at a ygaY site, a strong promoter Ptrc is used for controlling ilvIH (G14N S17F)-pykF to carry out synchronous multi-copy expression, and the valine production strain can be used for producing valine. On the basis of the strain, an artificial operon is constructed at an ycgH site, a strong promoter Ptrc is used for controlling ilvC-ilvD to carry out synchronous multi-copy expression, meanwhile, feedback inhibition of valine on ilvIH is relieved, and an ilvIH natural promoter is replaced by the strong promoter Ptrc, so that more pyruvic acid flows to valine; the constructed strain does not contain plasmids, has no defects, does not need induction, has the advantages of good genetic stability, high fermentation yield and the like, and can be used for efficiently synthesizing valine from the beginning by taking glucose as a substrate.

The invention belongs to the field of gene engineering, and particularly relates to a novel recombination system in aeromonas and application of the novel recombination system. A novel recombination system in the aeromonas disclosed by the invention contains a homologous recombination system expression vector, the expression vector expresses an operon under the control of an inducible promoter, and the operon contains three genes of a coding host specific exonuclease RedAahy, a single-stranded DNA annealing protein RedBahy and a single-stranded DNA binding protein RedSahy; the operon is derived from an aeromonas hydrophila (Aeromonas hydrophila) plasmid pR148, and is characterized in that the operon is derived from the aeromonas hydrophila (Aeromonas hydrophila) plasmid pR148. By utilizing the recombination system, efficient, simple and convenient gene insertion and deletion of aeromonas are successfully realized, and the application prospect is wide.

Disclosed are compositions, systems, kits, and methods for detecting an analyte in a sample by regulated in vitro transcription. The compositions, systems, kits, and methods typically comprise and / or utilize one or more components selected from: (a) an RNA polymerase; (b) an allosteric transcription factor (ATF), wherein the ATF binds an analyte or target molecule as a ligand; (c) a reporter molecule, and (d) an engineered transcription template; and / or any combination thereof. The engineered transcription template typically comprises a promoter sequence for the RNA polymerase and an operator sequence for the ATF. The promoter sequence and operator sequence are operably linked to a sequence encoding an RNA, wherein the ATF modulates transcription of the encoded RNA when the ATF binds the analyte or target molecule as a ligand, wherein the transcribed RNA generates or interferes with a detectable signal in conjunction with the reporter molecule.

The invention provides an L-lysine production strain as well as a strain construction method and application. A hom (V59A) is introduced on a genome by utilizing a homologous recombination gene editing technology to weaken a competitive approach of lysine; the lysC (T311I) which is not inhibited by L-lysine is overexpressed; the method comprises the following steps of: overexpressing lysE, lysA, ddh, dapB and asd, knocking out a lysI gene and multi-copying the lysE, carrying out synchronous multi-copying on the dapA and EcdapB genes through a manual operon P1676-EcdapB-Pa12-dapA, and carrying out synchronous multi-copying on the dapB, ddh, lysA and lysO genes through a manual operon Psod-dapB-ddh-Ptuf-lysO-lysA; the constructed strain does not contain plasmids, has no defects, does not need induction, has the advantages of good genetic stability, high fermentation yield and the like, and can be used for stably producing L-lysine.

The present invention relates to nucleic acids and modified immune cells or precursors thereof, comprising a first polynucleotide encoding an NF-κB p65-rTetO fusion protein comprising one or more transactivation domains (TAD) of human NF-κB p65 fused to a reverse Tet operator (rTetO) binding domain; a second polynucleotide comprising a Tet operator region (TetOR) for providing inducible expression of one or more transgene(s) operatively linked thereoto; and a third polynucleotide comprising a promoter directing transcription of p65-rTetO from a first strand of DNA, where the p65-rTetO is configured to induce transcription of a transgene of interest operatively linked to the TetOR in the presence of tetracycline or doxycycline (Dox) from an opposite strand of DNA.

The invention belongs to the technical field of biological gene engineering, and particularly relates to escherichia coli with high RNA content and a construction method and application thereof.A strong promoter is used for replacing a promoter of escherichia coli rDNA, an expression element of strong promoter + rDNA + terminator is constructed, plasmid is used as a carrier, the expression element is transferred into escherichia coli, and an escherichia coli engineering strain with high RNA yield is constructed; wherein the rDNA is derived from one or more of rrnA, rrnB, rrnC, rrnD, rrnE, rrnG and rrnH of rDNA operons of escherichia coli, and comprises 16S rDNA, 23S rDNA and 5S rDNA in the corresponding operons. By adopting the construction method, the RNA content of the escherichia coli can be obviously improved, and when the method is used for producing RNA, the RNA yield is obviously higher than that of an initial escherichia coli strain and common saccharomycetes for RNA production at present.

The invention relates to the technical field of microorganisms, and particularly discloses a construction method and application of a recombinant microorganism. According to the construction method of the recombinant microorganism, compared with an original strain, the expression quantity of threonine operon leader peptide of the recombinant microorganism is reduced, and the expression quantities of glyceraldehyde-3-phosphate dehydrogenase mutant, aspartate kinase / homoserine dehydrogenase I mutant, homoserine kinase and threonine synthase are increased; the glyceraldehyde-3-phosphate dehydrogenase mutant is characterized in that glycine at the 188th site in a glyceraldehyde-3-phosphate dehydrogenase protein sequence is mutated into serine; the aspartate kinase / homoserine dehydrogenase I mutant is characterized in that glycine at the 433rd site in an aspartate kinase / homoserine dehydrogenase I protein sequence is mutated into arginine; the starting strain is escherichia coli. According to the construction method disclosed by the invention, a novel recombinant microorganism with improved threonine yield can be obtained, and a novel method is provided for fermentation production of threonine.

The present disclosure relates to genetically modified E. coli host cells producing L-serine and expressing or overexpressing an operon comprising genes encoding a selected 3-phosphoglycerate dehydrogenase, a selected phosphoserine aminotransferase and a selected phosphoserine phosphatase, operably linked to selected Translation Initiation Regions (TIR) and operably linked to a selected promoter.

The invention discloses recombinant Escherichia coli for producing xylitol by efficiently utilizing L-arabinose without adding glycerol and application, and belongs to the technical field of genetic engineering. Escherichia coli with an araBAD operon and an L-xylulokinase gene lyxK knocked out is used as a starting strain, a glucose specific transporter gene ptsG and a D-xylulokinase gene xylB in a genome are knocked out, and a global regulatory factor CRP is mutated; on the basis of an araA, dpe and lxr three-enzyme expression universal method reported at present, the gene arrangement sequence and copy number in three-enzyme tandem expression are optimized, a promoter and an RBS sequence are optimized and screened, and three-enzyme genes are integrated to a specific position of an escherichia coli chromosome in an optimized multi-copy form. According to the method, the inhibition effect of glucose on L-arabinose is achieved, the in-vivo multienzyme catalysis efficiency of existing synthesis of xylitol from arabinose is remarkably improved, and the yield of precursor arabitol and the yield of the product xylitol are greatly improved.

A genetic construct comprises a promoter operably linked to a first coding sequence, which encodes tyrosine hydroxylase (TH), and a second coding sequence, which encodes GTP cyclohydrolase 1 (GCH1), wherein the second coding sequence is 3′ to the first coding sequence, and the first and second coding sequences are part of a single operon. The genetic construct does not encode aromatic amino acid decarboxylase (AADC).

The invention discloses an artificially synthesized corrinic acid operon, and the sequence of the artificially synthesized operon is obtained by integrating corrinic acid synthesis related genes and connecting and constructing the corrinic acid synthesis related genes in a reordering and end-to-end manner. According to the operon, a recombinant strain is synthesized on the basis of various chassis strains such as escherichia coli and rhodobacter sphaeroides, the corrinic acid and the derivatives thereof are synthesized through fermentation, the corrinic acid and the derivatives thereof can also be synthesized through a cell-free catalytic system, the problem that overlapping genes of a natural operon are difficult to modify is solved, and the synthesis efficiency and controllability of the corrinic acid are improved.

The invention discloses an Escherichia coli strain capable of bidirectionally regulating the copy number of CoIE1 type plasmids and application of the Escherichia coli strain. According to the method, escherichia coli Mach1-T1 is taken as an initial strain, a CRISPR / CAS9 gene editing technology is utilized, RNase E Q36R variant genes, rssB genes and pcnb genes are constructed into a polycistron expression cassette, and the polycistron expression cassette is placed under the control of an arabinose operon to replace endogenous pcnb genes of a genome. Each gene in the polycistron expression cassette is driven to translate by an independent ribosome binding site (RBS). By adjusting the concentration of the arabinose, continuous and bidirectional regulation and control of the copy number of the CoIE1 type plasmids from'inhibition 'to'overproduction' can be realized, and the method has important significance on industrial mass production of the CoIE1 type plasmids and diversified application of the CoIE1 type plasmids.

The present invention relates to a resistance-free plasmid production system. The present invention combines toxic transcripts / anti-toxic transcripts, operon-related elements and small RNAs to construct a plasmid comprising a nucleic acid fragment 3 encoding a small RNA, and host cell expression cassettes comprising a toxic gene expression cassette and an anti-toxic gene expression cassette, wherein the toxic gene expression cassette comprises a promoter and a nucleic acid fragment 1 for transcribing the toxic transcript, and the anti-toxic gene expression cassette comprises a promoter, an operon and a nucleic acid fragment 2 for transcribing the anti-toxic transcript. The reverse complementation of the nucleic acid fragment 3 and the nucleic acid fragment 1 produces a steric effect to inhibit the function of the toxic transcript, so that the entire plasmid production process does not need to be added with antibiotics or other inhibitors to realize the screening and production of the plasmid, thereby the harm of antibiotics and inhibitors to the plasmid in cell gene therapy is solved.

Disclosed herein are programmable bacteria for tumor-targeted immunotherapeutic delivery. In certain embodiments, the programmable bacteria comprise at least one synchronized lysis circuit contained in a single operon which are capable of being further engineered to cyclically produce anti-cancer therapeutic agents including but not limited to nanobodies against immune checkpoint inhibitors and over-expressed markers in cancers, toxins, tumor antigens, cytokines, and chemokines. In some embodiments, the programmable bacteria comprise at least one synchronized lysis circuit contained in a single operon and at least one plasmid producing a therapeutic agent, i.e., at least one plasmid comprising a nucleic acid sequence which encodes a therapeutic agent. The disclosure also provides methods of curing and treating cancer using the programmable bacteria.

The present disclosure describes genetically modified host cells expressing or overexpressing one or more genes of an L-serine pathway, or an operon comprising such one or more genes, wherein the genes or operon are operably linked to a constitutive promoter having a nucleotide sequence which is at least 70 %, such as at least 80%, such as at least 90%, such as at least 95%, such as at least 96%, such as at least 97%, such as at least 98%, such as at least 99%, such as 100% identical to the constitutive promoter comprised in anyone of SEQ ID NO: 1 to 16.

The invention belongs to the technical field of vitamin B2 production, and particularly relates to a preparation method of a safe high-yield strain for vitamin B2 production, which comprises the following steps: S1, strain source and screening: analyzing the current situation and development trend of vitamin B2, and screening the strain; s2, genetic engineering modification: evaluating the production performance and genetic stability of the strain, and carrying out mutagenesis treatment on the screened strain by using a chemical mutagenic agent to obtain a genetically mutated strain; s3, screening and identifying high-yield mutant strains: integrating the fragments containing the spore cereus dry riboflavin operon into a vitamin B2 genome to generate the mutant strains, and performing fine adjustment on the riboflavin operon in the mutant strains by utilizing a multiple genome editing system. And through technical popularization, the manufacturing cost of the product can be reduced, and the social benefit is obvious.

The invention discloses a high-induction-activity promoter derived from a tagatose operon of bacillus licheniformis, and belongs to the field of gene engineering. The promoter PfruK2 capable of being induced by a carbon source in a PTS (Pleurotropin Tyrosine Sequence) way is screened, the induced carbon substrate spectrum capable of being responded by the promoter is wide, the promoter is not affected by a glucose-mediated CCR effect, the induction intensity is high, and the induction peak value is improved by more than 50% compared with that of a quorum sensing strong promoter PLan.

The present disclosure relates to an L-arginine-producing microorganism in which the activity of a regulator of nitrate reductase operon expression is attenuated and the activity of glutamine synthetase is enhanced, and to a method for producing L-arginine using same, wherein the L-arginine-producing microorganism, in which the activity of the regulator of nitrate reductase operon expression is attenuated and the activity of glutamine synthetase is enhanced, has been confirmed to exhibit increased L-arginine production capability compared to a parent strain, and thus can be widely used for L-arginine production.

The invention belongs to the field of plant diseases, and particularly relates to a luminous labeled recombinant bacterium Xap-LUX and a construction method and application thereof. According to the present invention, a luxCDABE operon of a photobacterium phorbia luminescens is controlled by a constitutive promoter, and the luxCDABE operon is inserted into a chromosome of a peach bacterial perforation bacterium Xanthomonas arbocola pv.pruni so as to construct the stably expressed constitutive luminescence labeled recombinant bacterium Xap-LUX; the number of the bacteria can be evaluated conveniently, quickly and reliably based on the bioluminescence value of the bacteria. Therefore, the bioluminescence values of the luminescent labeled recombinant bacteria Xap-LUX in different medicaments are further quantitatively determined on a large scale, the antibacterial effects of different medicaments are evaluated, and the application of the strain and the reliability of the determination method are proved by using a conventional method.

The application discloses a method for identifying Klebsiella pneumoniae gene circadian rhythm expression by using a bioluminescence reporting system. The application constructs a recombinant bioluminescence Klebsiella pneumoniae driven by a metabolic gene malate:quinone oxidoreductase mqo promoter (Pmqo), and directly observes the change rule of bacterial gene expression under entrainment and free-running conditions of temperature cycle by monitoring the luminescence characteristics. The luxCDABE gene operon comprises genes for coding luciferase and substrate synthesis enzyme, wherein luxA and luxB code for heterodimer luciferase, luxC, luxD and luxE synthesize aldehyde as a substrate for the luciferase luminescence reaction. The luciferase catalyzes the oxidation of aldehyde to generate visible blue-green light with a wavelength of 450-490 nm. The Lux bioluminescence reporting system has the advantages of not needing to add an exogenous substrate and not affecting the normal physiological functions of the host, and can be applied to the identification of Klebsiella pneumoniae gene circadian rhythm expression.

Described herein are compositions and methods for generating a chassis for high-capacity production of different classes of high-value terpenoids. The chassis is a microorganism host that expresses a first operon comprising a modified ribosomal binding site, a Hmg-CoA reductase (HmgR) polypeptide, a 3-hydroxy-3-methylglutaryl CoA synthase (HmgS) polypeptide, and a β-ketothiolase (PhaA) polypeptide; and a second operon comprising a modified ribosomal binding site, a mevalonate kinase (MvK1) polypeptide, a phospho-mevalonate kinase (MvK2) polypeptide, and a diphosphomevalonate decarboxylase (MvD) polypeptide.

This invention provides an L-lysine-producing strain, its construction method, and its application. The strain utilizes homologous recombination gene editing technology to first overexpress... dapA , aspB Introducing mutations ppc (A482V) Knockout amn Multiple copies are synchronized by manually manipulating Pa12-dapB(T1)-Psod-aspB(T1)-Pa16-Ec_lysA. dapB , aspB Ec. from E. coli W3110 lysA Multiple copies are synchronized via the artificial manipulator Pa12-ddh(RBS)-lysE(T1)-Pa16-gapN(RBS)-amtB. ddh , lysE Streptococcus equi subsp. porphyria gapN , amtB The strain is plasmid-free, defect-free, requires no induction, and has advantages such as good genetic stability and high fermentation yield, enabling stable production of L-lysine.

The invention provides a CRISPR (clustered regularly interspaced short palindromic repeats) transposon gene integration system. The CRISPR transposon gene integration system comprises a vector pQCasTns containing transposase and Cas protein, and a vector pDoner containing a functional sequence, the pQCasTns comprises an operon and a T7 terminator which are connected in sequence; the pDoner comprises a functional sequence recognized by transposase and a crRNA sequence expression module of a targeted genome. The CRISPR transposon system provided by the invention solves a series of problems existing in genome editing of halomonas, and can realize multi-target knockout, high-efficiency deletion of long fragments and high-efficiency integration of exogenous genes.

The present application relates to a kind of genetically engineered bacteria of high-yield L-cysteine, construction method, and its application in the microbial fermentation preparation L-cysteine in L-cysteine. The present application is integrated by Esa quorum sensing system and lactose operon system, constructs a multifunctional QSI dynamic regulation system, realizes the coupling of target gene expression and cell growth, and is applied to the biosynthesis of L-cysteine, and obtains a high-yield L-cysteine of escherichia coli gene engineering strain. In addition, the QSI dynamic control system constructed in the present application dynamically controls lactose operon system using quorum sensing system, replaces the addition of traditional inducer IPTG, effectively saves the cost of industrial production and reduces the toxicity influence of inducer IPTG to cell, and has good production application value.

Disclosed herein are polynucleotides comprising sequences encoding coenzyme M synthase (ComF) linked to a heterologous regulatory element and methods of using the same. The polynucleotides may comprise synthetic operons comprising additional sequences encoding enzymes, e.g., a taurine-pyruvate aminotransferase, a sulfoacetaldehyde acetyl transferase, or a sulfopyruvate decarboxylase. Also disclosed herein are recombinant prokaryotic cells, e.g., recombinant bacterial, e.g., E. coli, or archaeal cells, e.g., Methanosarcina acetivorans with improved tolerance to oxidative stress.

The present disclosure relates to a genetically modified host cell that produces serine that expresses or overexpresses an operon comprising one or more genes encoding: a 3-phosphopyruvate dehydrogenase (serA) that converts D-3-phosphopyruvate (PGA) to phosphohydroxypyruvate (PHP); a phosphoserine transaminase (serC) for converting the 3-phosphohydroxypyruvic acid into phosphoserine, and a phosphoserine transaminase (serC) for the phosphoserine; and / or a phosphoserine phosphatase (serB) that converts phosphoserine to L-serine wherein the operon is operably linked to a constitutive promoter having a set of nucleotide sequences that are at least 95% (e.g., at least 96%, e.g., at least 97%, e.g., at least 98%, e.g., at least 99%, e.g., 100%) identical to the constitutive promoter comprised in SEQ ID NO: 8.