KR930021785A - Production of Human Proinsulin Fusion Proteins Using Genetic Manipulation - Google Patents
Production of Human Proinsulin Fusion Proteins Using Genetic Manipulation Download PDFInfo
- Publication number
- KR930021785A KR930021785A KR1019920005628A KR920005628A KR930021785A KR 930021785 A KR930021785 A KR 930021785A KR 1019920005628 A KR1019920005628 A KR 1019920005628A KR 920005628 A KR920005628 A KR 920005628A KR 930021785 A KR930021785 A KR 930021785A
- Authority
- KR
- South Korea
- Prior art keywords
- gene
- human proinsulin
- coli
- fusion protein
- proinsulin fusion
- Prior art date
Links
- 108010076181 Proinsulin Proteins 0.000 title claims abstract description 16
- 108020001507 fusion proteins Proteins 0.000 title claims abstract description 13
- 102000037865 fusion proteins Human genes 0.000 title claims abstract description 12
- 238000004519 manufacturing process Methods 0.000 title claims description 3
- 238000010353 genetic engineering Methods 0.000 title abstract 2
- 241000588724 Escherichia coli Species 0.000 claims abstract description 15
- 239000013604 expression vector Substances 0.000 claims abstract description 7
- 108090000623 proteins and genes Proteins 0.000 claims abstract 8
- 238000000034 method Methods 0.000 claims abstract 5
- 210000003705 ribosome Anatomy 0.000 claims abstract 4
- 150000001413 amino acids Chemical class 0.000 claims 1
- 230000002209 hydrophobic effect Effects 0.000 claims 1
- 102000004169 proteins and genes Human genes 0.000 claims 1
- 108700026220 vif Genes Proteins 0.000 claims 1
- 239000013598 vector Substances 0.000 abstract description 3
- 210000004027 cell Anatomy 0.000 abstract 1
- 238000010367 cloning Methods 0.000 abstract 1
- 239000002299 complementary DNA Substances 0.000 abstract 1
- 238000000855 fermentation Methods 0.000 abstract 1
- 230000004151 fermentation Effects 0.000 abstract 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 2
- 101001026869 Mus musculus F-box/LRR-repeat protein 3 Proteins 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
Abstract
본 발명은 유전자조작을 이용하여 사람의 프로인슐린 융합단백질을 생산하는 데 있어서 프로인슐린 융합단백질의 안정성과 발현율을 증가시키는 방법에 관한 것이다.The present invention relates to a method for increasing the stability and expression rate of proinsulin fusion proteins in producing human proinsulin fusion proteins using genetic engineering.
먼저 사람 프로인슐린 유전자를 cDNA 클로닝 방법으로 대장균 속주의 클로닝하였다. 클로닝된 사람 프로인슐린 유전자를 Lac 리보솜 결합부위 유전자, (Thr)6코딩하는 유전자와 함께 pUC8에 도입 [pUC8-(Thr)6-PI]하였고, pUC8-(Thr)6-PI와 pEX2로 터 pYK10-9 발현벡터를 제조하였다.First, the human proinsulin gene was cloned into the E. coli strain by the cDNA cloning method. The cloned human proinsulin gene was introduced into pUC8 with the Lac ribosomal binding site gene, (Thr) 6 encoding [pUC8- (Thr) 6- PI], and the pUC8- (Thr) 6- PI and pYK10 from pEX2 An -9 expression vector was prepared.
프로인슐린 전서열이 짧은 pYK10-9 벡터가 도입된 대장균에서도 발현되 융합 단백질은 대장균 세포내에서 안정하였다. pYK10-9 발현벡터가 형질전환된 대장균 pop2136은 플라스크 발효에서 249mg/1의 사람 프로인슐린 융합단백질을 생성하였고, 이 균을 유가배양한 결과 생성된 프로인슐린의 양은 2.2g/l로 현저히 증하하였다.The fusion protein was also expressed in E. coli cells in which Escherichia coli with a short pYK10-9 vector was introduced. Escherichia coli pop2136 transformed with pYK10-9 expression vector produced 249 mg / 1 human proinsulin fusion protein in flask fermentation.
Description
본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음Since this is an open matter, no full text was included.
제1도는 사람 프로인슐린 융합단백질 발현 벡터의 제작 개요도이다.1 is a schematic diagram of the production of human proinsulin fusion protein expression vectors.
제2도는 프로인슐린 유전자가 도입된 재조합 대장균으로부터 생성된 프로인슐린 융합단백질의 SDS-폴리아크릴마이드 젤 전기영동 사진이다.Figure 2 is a SDS-polyacrylamide gel electrophoresis picture of the proinsulin fusion protein produced from recombinant E. coli introduced with the proinsulin gene.
제3도는 재조합 벡터 pYK10-9가 도입된 대장균 pop2136이 플라스크 배양에서의 성장곡선이다.3 shows the growth curve of the Escherichia coli pop2136 into which the recombinant vector pYK10-9 was introduced.
제4도는 재조합 벡터 pYK10-9가 도입된 대장균 pop2136의 플라스크 배양에서 시간에 따른 발현단백질의 SDS-폴리아크릴아마이드 젤 전기 영동 사진이다.4 is a photograph of SDS-polyacrylamide gel electrophoresis of the expression protein over time in the flask culture of Escherichia coli pop2136 into which the recombinant vector pYK10-9 was introduced.
Claims (5)
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1019920005628A KR100221560B1 (en) | 1992-04-01 | 1992-04-01 | Production of insulin-fusion protein using genetic engineering |
| US07/954,364 US5460954A (en) | 1992-04-01 | 1992-09-30 | Production of human proinsulin using a novel vector system |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1019920005628A KR100221560B1 (en) | 1992-04-01 | 1992-04-01 | Production of insulin-fusion protein using genetic engineering |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR930021785A true KR930021785A (en) | 1993-11-23 |
| KR100221560B1 KR100221560B1 (en) | 1999-10-01 |
Family
ID=19331377
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1019920005628A KR100221560B1 (en) | 1992-04-01 | 1992-04-01 | Production of insulin-fusion protein using genetic engineering |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR100221560B1 (en) |
-
1992
- 1992-04-01 KR KR1019920005628A patent/KR100221560B1/en not_active IP Right Cessation
Also Published As
| Publication number | Publication date |
|---|---|
| KR100221560B1 (en) | 1999-10-01 |
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Payment date: 20060502 Year of fee payment: 8 |
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