97 results about "Coli strain" patented technology

The invention provides recombinant Escherichia coli. The recombinant Escherichia coli comprises the following multiple modifications: knocking out genes such as iclR, lpxM, ldhA, nadR, ptsP and the like; the expression of genes such as narJ, setA and the like is enhanced. The Escherichia coli disclosed by the invention can rapidly intake a glucose raw material, and can rapidly grow under various culture conditions. Therefore, the method has potential application value in the field of fermentation.

The invention discloses a biotin ligase mutant and application thereof, and belongs to the technical field of gene engineering. According to the invention, a series of biotin ligase mutants are prepared by docking biotin ligase and substrate molecules thereof and carrying out multiple mutations on screened 10 amino acids with binding force acting sites. On the basis, a biotin ligase mutant is connected to escherichia coli through a homologous recombination technology, a series of escherichia coli recombinant bacteria with high biotin yield are obtained, and the biotin yield reaches 0.873 mg / L to the maximum after 24-hour fermentation.

The invention discloses an anti-bacteriophage escherichia coli chassis cell based on combination of bacteriophage adaptive evolution and a bacterial anti-bacteriophage defense system. Receptor genes of bacteriophages are excavated through co-evolution of bacteriophages T1, T4, T7, vBEcoMP251 and vBEcoMP255 and escherichia coli strains, the resistance of escherichia coli to the bacteriophages is enhanced by knocking out screened bacteriophage receptors on the genome level, and meanwhile, defense systems shedu and septu with the ability to resist the bacteriophages are obtained through screening. The resistance of escherichia coli to the bacteriophage is further enhanced by integrating the gene to the ylbe and yeeP gene loci, construction of multi-resistant broad-spectrum chassis cells of the bacteriophage is completed, and certain application prospects are achieved.

The invention relates to a method for producing 6 '-fucosylated chitosan disaccharide GlcNacbeta1-4 (Fucalpha1-6) GlcNAc through double-bacterium coupling fermentation, and belongs to the technical field of fermentation engineering. The method comprises the following steps: taking fucose and N-acetyl-D-glucosamine as substrates in a fermentation system; the 6 '-fucosylated chitosan disaccharide is produced through coupled fermentation of an engineering escherichia coli strain and yeast, and a GlcNAcbeta1-4 (Fucalpha1-6) GlcNAc synthesis pathway including N-acetylhexosamine 1-kinase nahK, fucosyltransferase nodZ, L-fucose kinase Fkp and N, N-diacetylchitosan phosphorylase Chbp is constructed in the engineering escherichia coli, so that the 6'-fucosylated chitosan disaccharide is obtained. According to the invention, the yeast is added to realize cyclic regeneration from GMP to GTP, so that not only is the problem of high oligosaccharide production cost caused by pure exogenous supplement of GTP effectively solved, but also efficient production of GlcNAcbeta1-4 (Fucalpha1-6) GlcNAc is realized, and the yield of GlcNAcbeta1-4 (Fucalpha1-6) GlcNAc is increased to 25.50 g / L by an optimized coupling fermentation production technology. The invention provides a novel synthesis technology of complex N-glycans, and has a wide application prospect.

The invention discloses an escherichia coli engineering strain with high benzyl acetate yield and application thereof. The escherichia coli strain efficiently expresses a brand new biosynthetic pathway related gene of benzyl acetate; comprising a phenylalanine lyase gene, a cinnamic acid coenzyme A ligase gene, a 3-hydroxyacyl coenzyme A dehydrogenase gene, a 3-oxoacyl coenzyme A ketohydrolase gene, an enoyl coenzyme A hydratase gene, a carboxylic acid reductase gene, a phosphoric acid pantetheinyl transferase gene and an acyltransferase gene. The obtained escherichia coli engineering strain can utilize a sustainable and cheap raw material glucose as a carbon source to realize efficient synthesis of benzyl acetate. The yield of benzyl acetate synthesized by the escherichia coli benzyl acetate engineering bacteria is the highest yield of benzyl acetate synthesized by a microbial fermentation method reported at home and abroad from beginning. The benzyl acetate produced by the engineering strain has the advantages of reliable quality, economic product, green production and the like, and has important economic value and social benefit.

The invention relates to a diarrhea-related multi-drug-resistant coliphage and application thereof. In particular to a bacteriophage vBEcoMP66 screened from a multi-drug-resistant escherichia coli library, and the bacteriophage vBEcoMP66 is preserved in the China Center for Type Culture Collection on December 12, 2024, and the preservation number is CCTCC M 20242794. The bacillus subtilis has a splitting effect on multiple strains of multi-drug-resistant escherichia coli and piglet diarrhea pathogenic escherichia coli strains, and also has an obvious inhibition effect on bacteria in a biological membrane, and the inhibition capability is superior to the antibiotic level under the same condition. Besides, the bacteriophage has extremely low host resistance mutation frequency, the resistance mutation frequency of host bacteria can be effectively reduced, animal experiments show that the bacteriophage has the effects of preventing piglet diarrhea and reducing the level of inflammatory factors in vivo, and the bacteriophage has extremely high industrial value and wide application prospects.

The invention discloses a method for synthesizing alpha-arbutin through biotransformation, and belongs to the technical field of biochemistry. A gene cloning technology is utilized, sucrose phosphorylase Spase in leuconostoc mesenteroides and sucrose phosphorylase gtfa in bifidobacterium adolescentis are screened and found through a Deep Molecules and DLKcat algorithm on a Uniprot database, novopro is utilized for codon optimization, then the sucrose phosphorylase Spase and the sucrose phosphorylase gtfa are expressed on a plasmid vector PET-28a and are converted into an escherichia coli BL21 (DE3) strain, and the sucrose phosphorylase Spase and the sucrose phosphorylase gtfa in the bifidobacterium adolescentis are obtained. An engineering strain for heterologous expression of the gene is constructed by taking the gene as a chassis cell. According to the process, sucrose and hydroquinone are taken as co-substrates, and bioconversion is carried out under the action of sucrose phosphorylase from different sources to obtain alpha-arbutin.

The invention provides a bacterial outer membrane vesicle drug carrier with a targeting effect and preparation, and relates to the technical field of biological medicines.The bacterial outer membrane vesicle protein ClyA derived from an escherichia coli W3110 strain is combined with iRGD protein capable of being combined with an integrin receptor alpha v beta 3, a ClyA-iRGD-pGEX-6P-1 recombinant plasmid for expressing ClyA-iRGD fusion protein is constructed, and the ClyA-iRGD-pGEX-6P-1 recombinant plasmid is used for preparing the bacterial outer membrane vesicle drug carrier with the targeting effect. By endogenous expression of the plasmid in W3110 escherichia coli, the C-iRGD-OMVs which has a tumor targeting effect and shows ClyA-iRGD fusion protein on the surface is prepared. The C-iRGD-OMVs shows efficient targeting binding capacity to MCF-7 breast cancer cells, overcomes the defect that iRGD single peptide is prone to aggregation and crystallization, has the potential of loading drugs, lays a foundation for the C-iRGD-OMVs serving as a targeting carrier for delivering breast cancer treatment drugs, and is expected to be further developed into a targeting treatment preparation for breast cancer.

The invention belongs to the technical field of biology, and relates to a zero-background carrier as well as a preparation method and application thereof. The zero-background vector provided by the invention comprises an optimized escherichia coli CcdB gene and a corresponding escherichia coli promoter, and the nucleotide sequence of the zero-background vector is shown as SEQ ID NO.1. The zero background vector is applicable to any plasmid vector; both a cloning vector plasmid and an expression vector plasmid have relatively high connection efficiency and positive rate, and a connection product can be directly converted into a common escherichia coli strain, so that a target gene cloning vector can be effectively prepared; the target plasmid inserted with the zero background carrier has high short fragment insertion efficiency, and can also be used for long fragment insertion at the same time. In the genome assembly process, no matter whether a small fragment or a large gene fragment is inserted, the method can ensure an efficient assembly effect, especially in small fragment gene cloning, the efficiency and the positive rate can be remarkably improved, and meanwhile generation of false positive cloning is reduced.

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 provides anti-escherichia coli endolysin E1, E2 and E3 and application thereof. Specifically, the invention discloses preparation and application of three anti-escherichia coli endolysin E1, E2 and E3. Through a gene engineering technology, E1, E2 and E3 are seamlessly cloned to a pSumo-mut vector respectively, and are subjected to prokaryotic expression in an escherichia coli BL21 (DE3) strain. Through antibacterial activity evaluation, when the endolysin E1, the endolysin E2 and the endolysin E3 are independently used or cooperatively used, the endolysin E1, the endolysin E2 and the endolysin E3 show an obvious bactericidal effect on escherichia coli isolates, particularly on multi-drug-resistant clinical isolates. The invention provides a new thought for developing a novel anti-escherichia coli drug, and has a good potential value in the aspect of developing an anti-escherichia coli infection drug.

The invention relates to the technical field of biological medicines, in particular to an Escherichia coli strain, a pharmaceutical composition and application, and more particularly relates to tumor inhibitory Escherichia coli 08 and application thereof. The preservation number of the escherichia coli strain is CCTCC (China Center For Type Culture Collection) NO: M20251869. After tumor cells are infected by the escherichia coli strain, the tumor microenvironment can be changed, and the tumor cells are damaged, so that the activity of the tumor cells is effectively inhibited.

This disclosure provides an E. coli strain, which lacks thioredoxin reductase activity encoded by trxB and thioredoxin 1 activity encoded by trxA, and glutathione reductase activity encoded by gor. Said E. coli strain expresses a mutated AhpC protein having glutathione reductase activity and a cytosolic prokaryotic disulfide isomerase. The E. coli strain has an oxidative cytosol and can be used to efficiently produce proteins having disulfide bonds.

The invention discloses an escherichia coli system for synthesizing D-lactic acid as well as a preparation method and application of the escherichia coli system. The escherichia coli system comprises a strain GL10 and a strain XL15; the bacterial strain GL10 is obtained by deleting genes of ppsA, mgsA, dld, pta, poxB, pflB, adhE, frdA, frdB and frdC in an escherichia coli BL21 bacterial strain; the bacterial strain XL15 is obtained by deleting genes of glk, ptsI, ptsG, mglB and galP in a bacterial strain GL10. The strain GL10 efficiently utilizes glucose, and the strain XL15 utilizes xylose, so that the CCR effect is effectively relieved, efficient synergistic utilization of a mixed carbon source is realized, the yield and purity of D-lactic acid are improved, and the production cost is reduced.

The invention discloses a synthesis method of ubiquitin (1-74) NHNH2, and belongs to the technical field of protein synthesis. The synthesis method comprises the following steps: utilizing a prokaryotic expression system, expressing a full-length protein of Ub (1-76) and a Josephin protein coded by Burkholderia pyrrocinia by virtue of an escherichia coli BL21 (DE3) strain, hydrolyzing the Ub (1-76) by virtue of the obtained 6His-BpJOS, cutting a peptide bond between Ub Arg-74 and Gly-75, and capturing a generated thiolipid intermediate by virtue of hydrazine hydrate, so as to obtain the Ub (1-74) NHNH2. According to the present invention, the ubiquitin cleavage enzyme BpJOS catalyzed in-situ activation process is adopted, such that the preparation yield is high, the operation is simple, the large-scale preparation can be achieved, the synthesis cost is low, and the method can be used for synthesizing the multi-ubiquitination modified fragment and synthesizing the Ub (1-74) probe as the precursor, and has wide application prospects.

The invention discloses an escherichia coli strain with high yield of riboflavin as well as a construction method and application of the escherichia coli strain. A series of Escherichia coli strains with high yield of riboflavin are constructed, the constructed Escherichia coli with high yield of riboflavin is safe and harmless, glucose is used as a substrate for fermentation under the aerobic condition of a shake flask, the yield of riboflavin is 1117.8 mg / L, 1330.1 mg / L, 1430.8 mg / L, 1508.7 mg / L and 1640.8 mg / L in sequence, and the yields are 112.5 mg / g of glucose, 134.1 mg / g of glucose, 146.3 mg / g of glucose, 158.8 mg / g of glucose and 168.1 mg / g of glucose in sequence; when the strain SN04M is subjected to fed-batch fermentation in a 1L fermentation tank, the yield of riboflavin reaches 8.52 g / L, and the total yield is 189.9 mg / g glucose. According to the invention, the fermentation yield of riboflavin and the substrate utilization rate of riboflavin to glucose can be obviously improved.

The present disclosure provides materials and assays, herein referred to as surface display ACE (surfACE), that use E. coli strains expressing, in one embodiment, nanobodies (VHHs) on the outer cell membrane as a solution to increase de novo screening throughput and efficiency using a combination of MACS and FACS based enrichment strategies.

Disclosed are engineered E. coli Nissle strains having increased transformation efficiency. Also described are methods for generating transformants using the strains. Probiotic compositions containing the engineered E. coli Nissle strains and methods of using the probiotic compositions are also described herein.

The invention discloses an Escherichia coli strain capable of stably expressing near-infrared fluorescent protein as well as a construction method and application of the Escherichia coli strain, and belongs to the technical field of gene engineering. According to the bacterial strain, escherichia coli probiotics Nissle 1917 serve as a chassis bacterial strain, at least two miRFP713 gene expression cassettes are integrated on a chromosome through a CRISPR-Cas mediated homologous recombination technology, and each expression cassette comprises a constitutive promoter and a miRFP713 gene optimized by a codon; meanwhile, the biliverdin synthetase gene is integrated to realize cofactor self-supply. The chromosome integrative Escherichia coli strain constructed by the invention realizes stable expression without exogenous induction, the fluorescence signal retention rate of 20 generations of passage is greater than 98%, and the fluorescence intensity is improved by 94.8 times, and is suitable for multiple application scenes such as long-term living body tracking, tumor targeted tracking and the like.

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 invention provides Escherichia coli for producing a nylon 12 monomer. The genotypes of the Escherichia coli are as follows: tesA: PtluxR-PluxluxIUcfatB, adhE: PtluxR-PluxluxIUcfatB, fadE: PluxCYP153A-BM3, pgi: PluxCYP153A-BM3, aldB: PluxCYP153A-BM3, and gabD: PM12CV2025BsADH. The invention also provides a preparation method of the Escherichia coli. The genotypes of the Escherichia coli are as follows: tetluxR: PtluxR: PluxluxIUcfatB, adhE: The invention further provides a construction method of the escherichia coli for producing the nylon 12 monomer, a self-induction system in the escherichia coli is constructed by utilizing a Lu xI / R quorum sensing system in vibrio fischeri, and the synthesis rate of AHL is slowed down, so that the self-induction expression of the gene is realized. An engineering escherichia coli strain is constructed based on an established self-induction system, self-induction synthesis of the nylon 12 monomer can be realized, cell growth and product synthesis are balanced, and synthesis of the nylon 12 monomer from glucose is realized.

The invention discloses Escherichia coli for fermentation production of riboflavin anti-bacteriophage, and belongs to the technical field of microorganisms. Receptor genes of bacteriophages are excavated through co-evolution of bacteriophages T1, T7, vBEcoMP251 and vBEcoMP255 and escherichia coli strains, the resistance of escherichia coli to the bacteriophages is enhanced by knocking out screened bacteriophage receptors on the genome level, and meanwhile, defense systems Gabija and KELM with the ability of resisting bacteriophages JNUWH1 and JNUWD and defense systems Shedu and Septu with the ability of resisting bacteriophages T4 are obtained through screening. Gene is integrated to a genome to strengthen the resistance of escherichia coli to bacteriophage, construction of the anti-bacteriophage chassis cell is completed, meanwhile, genes lacI, pfkA, edd and eda are knocked out, the anti-bacteriophage escherichia coli chassis cell for fermentation production of riboflavin is obtained, and a certain application prospect is achieved.

The invention belongs to the technical field of genetic engineering, and discloses a genetically engineered bacterium for efficiently synthesizing gamma-aminobutyric acid from the beginning by efficiently utilizing a mixed carbon source, a method and application, and the genetically engineered bacterium is obtained by overexpressing edd from Zymomonas mobilis on the basis of escherichia coli E. coli G16; eda from the zymomonas mobilis is over-expressed; performing overexpression on acs (Acetobacter pasteurianus) sources; a malic acid synthase gene aceB is knocked out; a malic acid synthase gene glcB is knocked out; and the acetyl phosphate transferase gene eutD is knocked out. The escherichia coli strain which is clear in genetic background and capable of producing gamma-aminobutyric acid is utilized, and an ED path and an acetic acid utilization path are introduced, so that acetyl CoA branch metabolism is reduced, and the yield and conversion rate of gamma-aminobutyric acid are remarkably improved.

The invention discloses a high-concentration malonic acid stress tolerant escherichia coli chassis cell and a construction method thereof. The construction method of the high-concentration malonic acid stress tolerant escherichia coli chassis cell comprises the following steps: carrying out high-concentration malonic acid tolerance domestication on an escherichia coli strain ZXP05 by utilizing an adaptive laboratory evolution (ALE) technology to obtain a high-concentration malonic acid tolerant escherichia coli strain. The strain is subjected to genome re-sequencing, and the result shows that G at the 192nd site from the 5'tail end of the nucleotide sequence of the ompR gene on the genome of the tolerant strain is mutated into C. And mutating G at the 192 site from the 5'tail end of the nucleotide sequence of the ompR gene into C in a homologous recombination manner for a wild escherichia coli strain E.coli C43 (DE3), thereby successfully obtaining the escherichia coli strain W-TB capable of tolerating 400 mM malonic acid. The construction method disclosed by the invention is high in universality, simple and effective, and high osmotic pressure stress of the strain can be relieved; the finally obtained strain W-TB can tolerate high-concentration malonic acid, and the strain can be used as a chassis cell to be applied to biosynthesis of high-added-value compounds by taking high-concentration malonic acid as a substrate and biosynthesis of malonic acid.

The invention provides an Escherichia coli strain for efficiently synthesizing ergothioneine and application of the Escherichia coli strain to fermentation production of ergothioneine. Specifically, the invention provides an escherichia coli strain E1 for expressing high-activity ergothioneine synthetase 1 (cpEGT1-M) and ergothioneine synthetase 2 (clEGT2-M), and 8.6 g / L of ergothioneine is obtained through fermentation. Meanwhile, high-activity S-adenosylmethionine synthetase ctMetK-M is introduced into the strain E1, the synthesis step of S-adenosylmethionine playing a speed limiting role in the strain E1 is accelerated, an escherichia coli strain E2 is obtained, and 15.3 g / L of ergothioneine is obtained through fermentation. Furthermore, high-activity glutathione reductase pfGR-M is introduced into the strain E2, an escherichia coli strain E3 is obtained, and 20.7 g / L of ergothioneine is obtained through fermentation.

The invention provides a genetic engineering strain for efficiently producing beta-alanine, an escherichia coli strain is modified by adopting a genetic engineering means, and a multi-plasmid system is constructed, so that a metabolic pathway is optimized and the product synthesis efficiency is improved. The method comprises the following steps: firstly, by constructing a recombinant plasmid pET-PanD, introducing aspartic acid decarboxylase (L17F / G24R mutant) with mutation so as to promote efficient conversion of aspartic acid to beta-alanine; meanwhile, phosphoenolpyruvate carboxylase (PPC) is overexpressed through the recombinant plasmid pRSF-PPC, generation of oxaloacetic acid (OAA) is increased, and then the synthesis path of carbon flow to beta-alanine is improved. Besides, NADH dependent type aspartate dehydrogenase (AspDH) from pseudomonas aeruginosa is overexpressed by adopting a pBAD-AspDH plasmid, so that the dependence on NADPH is reduced, and the conversion efficiency is further improved.

The invention provides an sgRNA molecule of a targeted IMP-4 gene and a biomacromolecular compound containing the sgRNA molecule, the biomacromolecular compound forms a CRISPR-Cas3 system for targeted removal of the IMP-4 drug-resistant gene, and the invention also provides a method for targeted removal of the IMP-4 drug-resistant gene in a strain by using the system. According to the CRISPR-Cas3 system provided by the invention, the directed gene clearance rate of the strain carrying the IMP-4 drug-resistant gene can reach 100%. The minimum inhibitory concentration of the Escherichia coli subjected to targeted removal of IMP-4 to meropenem is averagely reduced by 71.11 times compared with that of a strain without targeted removal of IMP-4.

The present invention discloses a method for modifying Escherichia coli to produce phycocyanin, belonging to the fields of genetic engineering and bioengineering. The method heterologously expresses the ho1 and pcyA genes from Synechocystis sp. PCC6803, converting protoheme into biliverdin, an intermediate in phycocyanin synthesis, while reducing the accumulation of biliverdin, an intermediate in phycocyanin synthesis. Furthermore, the E. coli genome is modified to overexpress genes involved in the phycocyanin metabolic pathway, resulting in a recombinant E. coli strain with a high phycocyanin production. When the recombinant strain is cultured for 36 hours in a system using inexpensive glycerol as a substrate, the phycocyanin yield reaches 147 mg / L.

The present invention relates to a Ses i 2-specific binding polypeptide of sesame allergen and a screening method thereof. The specific binding polypeptide includes the polypeptides shown in SEQ ID NO.1-5. The method includes the following steps: (I) Activate and culture Escherichia coli strain 2738, add phages with gradient dilution to obtain a recombinant phage library; (II) Perform three rounds of panning with Ses i 2 protein as the target, amplify the bound target phages, sequence them with 96gIII sequencing primers, and obtain a polypeptide specifically binding to Ses i 2 protein after translation; (III) Synthesize the polypeptide obtained in the above steps and verify the affinity by bio-layer interferometry. The screening method uses a phage library to screen for peptide aptamers of sesame protein Sesi 2, which have high affinity and specificity and can be used for the detection of Ses i 2 protein.