scholarly journals Clinical manufacturing of recombinant human interleukin 15. I. Production cell line development and protein expression in E. coli with stop codon optimization

2011 ◽  
Vol 28 (2) ◽  
pp. 497-507 ◽  
Author(s):  
Vinay V. Vyas ◽  
Dominic Esposito ◽  
Terry L. Sumpter ◽  
Trevor L. Broadt ◽  
James Hartley ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cell Line ◽  
Codon Optimization ◽  
Stop Codon ◽  
Cell Line Development ◽  
Production Cell Line ◽  
E Coli ◽  
Interleukin 15 ◽  
Production Cell

Codon Optimization Enhances Protein Expression of Bombyx mori Nucleopolyhedrovirus DNA Polymerase in E. coli

2013 ◽  
Vol 68 (3) ◽  
pp. 293-300 ◽  
Author(s):  
Huifang Song ◽  
Guohui Li ◽  
Weijun Mai ◽  
Guoping Huang ◽  
Keping Chen ◽  
...  
Keyword(s):  
Protein Expression ◽  
Bombyx Mori ◽  
Dna Polymerase ◽  
Codon Optimization ◽  
Bombyx Mori Nucleopolyhedrovirus ◽  
E Coli

scholarly journals Strategy for Designing the Synthetic Gene Encoding Human papillomavirus Major Capsid L1 Protein for Heterologous Expression in Escherichia coli System

2020 ◽  
Vol 8 (2) ◽  
pp. 225
Author(s):  
Kartika Sari Dewi ◽  
Wien Kusharyoto
Keyword(s):  
Escherichia Coli ◽  
Protein Expression ◽  
Codon Optimization ◽  
Gc Content ◽  
Web Server ◽  
Synthetic Gene ◽  
Gene Encoding ◽  
E Coli ◽  
Gene Construct ◽  
L1 Protein

DNA is widely used to construct heterologously expressed genes. The adaptation of the codons to the host organism is necessary in order to ensure sufficient production of proteins. The GC content, codon identity and the mRNA from the translation site are also important in the design of the gene construct. This study performed a strategy for the design of synthetic gene encoding HPV52 L1 protein and several analyses at the genetic level to optimize its protein expression in the Escherichia coli BL21(DE3) host. The determination of the codon optimization was performed by collecting 75 HPV52 L1 protein sequences in the NCBI database. Furthermore, all the sequences were analyzed using multiple global alignments by Clustal Omega web server. Once the model was determined, codon optimization was performed using OPTIMIZER and the web server of the IDT codon optimization tool based on the E. Coli B. The generated open reading frame (ORF) sequence was analyzed using Restriction mapper web server to choose the restriction site for facilitating the cloning stage, which is adjusted for pJExpress414 expression vector. To maximize the protein expression level, the mRNA secondary structure analysis around the ribosome binding site (rbs) was performed. A slight modification at the 5’-terminal end waa carried out in order to get more accessible rbs and increasing mRNA folding free energy. Finally, the construction of the synthetic gene was confirmed to ensure that no mutation occurs in the protein and to calculate its Codon Adaptation Index (CAI) and GC content. The above strategy, which leads to a good ORF sequence with the value of the free mRNA folding energy around rbs, is -5.5 kcal / mol, CAI = 0.787 and GC content 49.5%. This result is much better than its original gene. This result is much better compared to its native gene. Theoretically it is possible that this synthetic gene construct generates a high level protein expression in E. coli BL21 (DE3) under the regulation of the T7 promoter.

Response of a CHO production cell line to different types of hydrodynamic shear stress present in stirred and sparged bioreactors

2012 ◽  
Vol 29 ◽  
pp. S165
Author(s):  
Jochen B. Sieck ◽  
Thomas Villiger ◽  
Wolfgang E. Budach ◽  
Zoltan Suemeghy ◽  
Christian Leist ◽  
...  
Keyword(s):  
Shear Stress ◽  
Cell Line ◽  
Production Cell Line ◽  
Different Types ◽  
Hydrodynamic Shear ◽  
Production Cell

scholarly journals Physical methods for synchronization of a human production cell line

2011 ◽  
Vol 5 (Suppl 8) ◽  
pp. P49 ◽  
Author(s):  
Oscar Platas Barradas ◽  
Uwe Jandt ◽  
Ralf Hass ◽  
Cornelia Kasper ◽  
Volker Sandig ◽  
...  
Keyword(s):  
Cell Line ◽  
Production Cell Line ◽  
Physical Methods ◽  
Production Cell

Expression of GnTIII in a recombinant anti-CD20 CHO production cell line: Expression of antibodies with altered glycoforms leads to an increase in ADCC through higher affinity for FC?RIII

2001 ◽  
Vol 74 (4) ◽  
pp. 288-294 ◽  
Author(s):  
Julian Davies ◽  
LiYing Jiang ◽  
Li-Zhen Pan ◽  
Michael J. LaBarre ◽  
Darrell Anderson ◽  
...  
Keyword(s):  
Cell Line ◽  
Production Cell Line ◽  
Anti Cd20 ◽  
Production Cell

scholarly journals Codon Preference Optimization Increases Prokaryotic Cystatin C Expression

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Qing Wang ◽  
Cui Mei ◽  
Honghua Zhen ◽  
Jess Zhu
Keyword(s):  
Protein Expression ◽  
Cystatin C ◽  
Production Systems ◽  
Codon Optimization ◽  
Recombinant Protein Expression ◽  
Expression System ◽  
Gc Content ◽  
Optimization Techniques ◽  
E Coli ◽  
Prokaryotic Expression Vector

Gene expression is closely related to optimal vector-host system pairing in many prokaryotes. Redesign of the humancystatin C(cysC) gene using the preferred codons of the prokaryotic system may significantly increasecysCexpression inEscherichia coli(E. coli). Specifically,cysCexpression may be increased by removing unstable sequences and optimizing GC content. According toE. coliexpression system codon preferences, the gene sequence was optimized while the amino acid sequence was maintained. The codon-optimizedcysC(co-cysC) and wild-typecysC(wt-cysC) were expressed by cloning the genes into a pET-30a plasmid, thus transforming the recombinant plasmid intoE. coliBL21. Before and after the optimization process, the prokaryotic expression vector and host bacteria were examined for protein expression and biological activation of CysC. The recombinant proteins in the lysate of the transformed bacteria were purified using Ni2+-NTA resin. Recombinant protein expression increased from 10% to 46% based on total protein expression after codon optimization. Recombinant CysC purity was above 95%. The significant increase incysCexpression inE. coliexpression produced by codon optimization techniques may be applicable to commercial production systems.

Mapping and partial characterization of proteases expressed by a CHO production cell line

2006 ◽  
Vol 95 (5) ◽  
pp. 961-971 ◽  
Author(s):  
H. Sandberg ◽  
D. Lütkemeyer ◽  
S. Kuprin ◽  
M. Wrangel ◽  
A. Almstedt ◽  
...  
Keyword(s):  
Cell Line ◽  
Partial Characterization ◽  
Production Cell Line ◽  
Production Cell
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