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Engineering selection stringency on expression vector for the production of recombinant human alpha1-antitrypsin using Chinese Hamster ovary cells

Overview of attention for article published in BMC Biotechnology, June 2015
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  • In the top 25% of all research outputs scored by Altmetric
  • High Attention Score compared to outputs of the same age (83rd percentile)
  • Average Attention Score compared to outputs of the same age and source

Mentioned by

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1 news outlet
twitter
1 tweeter

Citations

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28 Dimensions

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52 Mendeley
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Title
Engineering selection stringency on expression vector for the production of recombinant human alpha1-antitrypsin using Chinese Hamster ovary cells
Published in
BMC Biotechnology, June 2015
DOI 10.1186/s12896-015-0145-9
Pubmed ID
Authors

Christine Lin Chin, Hing Kah Chin, Cara Sze Hui Chin, Ethan Tingfeng Lai, Say Kong Ng

Abstract

Expression vector engineering technology is one of the most convenient and timely method for cell line development to meet the rising demand of novel production cell line with high productivity. Destabilization of dihydrofolate reductase (dhfr) selection marker by addition of AU-rich elements and murine ornithine decarboxylase PEST region was previously shown to improve the specific productivities of recombinant human interferon gamma in CHO-DG44 cells. In this study, we evaluated novel combinations of engineered motifs for further selection marker attenuation to improve recombinant human alpha-1-antitrypsin (rhA1AT) production. Motifs tested include tandem PEST elements to promote protein degradation, internal ribosome entry site (IRES) mutations to impede translation initiation, and codon-deoptimized dhfr selection marker to reduce translation efficiency. After a 2-step methotrexate (MTX) amplification to 50 nM that took less than 3 months, the expression vector with IRES point mutation and dhfr-PEST gave a maximum titer of 1.05 g/l with the top producer cell pool. Further MTX amplification to 300 nM MTX gave a maximum titer of 1.15 g/l. Relative transcript copy numbers and dhfr protein expression in the cell pools were also analysed to demonstrate that the transcription of rhA1AT and dhfr genes were correlated due to the IRES linkage, and that the strategies of further attenuating dhfr protein expression with the use of a mutated IRES and tandem PEST, but not codon deoptimization, were effective in reducing dhfr protein levels in suspension serum free culture. Novel combinations of engineered motifs for further selection marker attenuation were studied to result in the highest reported recombinant protein titer to our knowledge in shake flask batch culture of stable mammalian cell pools at 1.15 g/l, highlighting applicability of expression vector optimization in generating high producing stable cells essential for recombinant protein therapeutics production. Our results also suggest that codon usage of the selection marker should be considered for applications that may involve gene amplification and serum free suspension culture, since the overall codon usage and thus the general expression and regulation of host cell proteins may be affected in the surviving cells.

Twitter Demographics

The data shown below were collected from the profile of 1 tweeter who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 52 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Mexico 1 2%
Denmark 1 2%
Canada 1 2%
Slovenia 1 2%
Unknown 48 92%

Demographic breakdown

Readers by professional status Count As %
Researcher 15 29%
Student > Ph. D. Student 12 23%
Student > Bachelor 5 10%
Other 3 6%
Student > Postgraduate 3 6%
Other 5 10%
Unknown 9 17%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 24 46%
Agricultural and Biological Sciences 12 23%
Engineering 2 4%
Unspecified 1 2%
Nursing and Health Professions 1 2%
Other 4 8%
Unknown 8 15%

Attention Score in Context

This research output has an Altmetric Attention Score of 10. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 03 June 2015.
All research outputs
#557,282
of 5,180,928 outputs
Outputs from BMC Biotechnology
#74
of 414 outputs
Outputs of similar age
#28,461
of 175,004 outputs
Outputs of similar age from BMC Biotechnology
#23
of 38 outputs
Altmetric has tracked 5,180,928 research outputs across all sources so far. Compared to these this one has done well and is in the 89th percentile: it's in the top 25% of all research outputs ever tracked by Altmetric.
So far Altmetric has tracked 414 research outputs from this source. They receive a mean Attention Score of 4.8. This one has done well, scoring higher than 79% of its peers.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 175,004 tracked outputs that were published within six weeks on either side of this one in any source. This one has done well, scoring higher than 83% of its contemporaries.
We're also able to compare this research output to 38 others from the same source and published within six weeks on either side of this one. This one is in the 36th percentile – i.e., 36% of its contemporaries scored the same or lower than it.