http://rdf.ncbi.nlm.nih.gov/pubchem/patent/CN-111065738-A
Outgoing Links
| Predicate | Object |
|---|---|
| assignee | http://rdf.ncbi.nlm.nih.gov/pubchem/patentassignee/MD5_261b513336bb6b507d1f313c6f0da3d1 |
| classificationCPCAdditional | http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/Y02A50-30 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12R2001-19 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N1-205 |
| classificationCPCInventive | http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/A61K48-00 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N15-00 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N1-205 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N9-14 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N15-69 http://rdf.ncbi.nlm.nih.gov/pubchem/patentcpc/C12N15-70 |
| classificationIPCInventive | http://rdf.ncbi.nlm.nih.gov/pubchem/patentipc/C12N15-90 http://rdf.ncbi.nlm.nih.gov/pubchem/patentipc/C12N15-63 http://rdf.ncbi.nlm.nih.gov/pubchem/patentipc/C12N5-10 http://rdf.ncbi.nlm.nih.gov/pubchem/patentipc/A61K48-00 http://rdf.ncbi.nlm.nih.gov/pubchem/patentipc/C12N15-09 |
| filingDate | 2018-03-26-04:00^^<http://www.w3.org/2001/XMLSchema#date> |
| inventor | http://rdf.ncbi.nlm.nih.gov/pubchem/patentinventor/MD5_289aebb59aa3391ecdc302ee972035fc |
| publicationDate | 2020-04-24-04:00^^<http://www.w3.org/2001/XMLSchema#date> |
| publicationNumber | CN-111065738-A |
| titleOfInvention | Gene therapy DNA vector VTvaf17, production method; Escherichia coli strain SCS110-AF, production method; Escherichia coli strain SCS110-AF/VTvaf17 carrying gene therapy DNA vector VTvaf17, production method |
| abstract | The present invention relates to genetic engineering and has applications in biotechnology, medicine and agriculture. The present invention requires the construction of a 3165bp gene therapy DNA vector VTvaf17 for genetic modification of animal and human cells containing the nucleotide sequence of SEQ ID No. 1. The method for constructing the 3165bp gene therapy DNA vector VTvaf17 involves first constructing a 4182bp vector containing 1188bp of human elongation factor EF1A with an intrinsic enhancer, then cutting it through the SpeI restriction site, and ligating the remaining fragment to itself Promoter region, 35 bp polylinker with sites for restriction endonucleases BamHI, EcoRV, Sail, HindIII, KpnI, EcoRI, 466 bp transcription terminator and polyadenylation sequence for human growth factor, allows positive without antibiotics 136bp regulatory element RNA-OUT of selected transposon Tn10, 1299bp origin of replication for autonomous replication with single nucleotide substitutions to increase vector generation in cells of most E. coli strains, 1010bp kanamycin resistance gene . The stated purpose can be achieved by obtaining the E. coli strain SCS110‑AF to generate a gene therapy DNA vector VTvaf17 or a gene therapy DNA vector based thereon that allows positive selection without antibiotics. The method for obtaining E. coli strain SCS110‑AF to generate the gene therapy DNA vector VTvaf17 or gene therapy DNA vectors based thereon involves the construction of a linear DNA fragment of 64 bp, followed by electroporation to transform E. A clone surviving in culture medium containing antibiotics, the linear DNA fragment containing the regulatory element RNA-IN of the transposon Tn10 allowing positive selection without antibiotics, the 1422 bp fructansucrase gene sacB (the product of which is ensured in cultures containing sucrose) The 763bp chloramphenicol resistance gene catR required for homologous recombination clones, two homologous sequences 329bp and 233bp that ensure homologous recombination in the recA region of the gene at the same time as gene inactivation . Escherichia coli strain SCS110‑AF/VTvaf17 carrying the gene therapy DNA vector VTvaf17 (registered at the Russian State Collection of Industrial Microorganisms under number B‑12990, International Depositary Agency number NCIMB 42801) was also constructed for use in allowing antibiotic-free selection. its further development. A method for obtaining the E. coli strain SCS110‑AFA/Tvaf17 (registered in the Russian State Collection of Industrial Microorganisms under the number B‑12990, the International Depositary Agency number NCIMB 42801) carrying the gene therapy DNA vector VTvaf17 comprising the preparation of a electrocompetent cells and electroporated with the gene therapy DNA vector VTvaf17. Afterwards, cells were poured into agar plates (Petri dishes) with selective medium containing yeast extract, peptone, 6% sucrose and 10 μg/ml chloramphenicol. |
| priorityDate | 2017-08-25-04:00^^<http://www.w3.org/2001/XMLSchema#date> |
| type | http://data.epo.org/linked-data/def/patent/Publication |
Incoming Links
Total number of triples: 620.