Abstract:
:Targeted gene integration into site-specific loci can be achieved in Chinese hamster ovary (CHO) cells via CRISPR/Cas9 genome editing technology and the homology-directed repair (HDR) pathway. The low efficiency of HDR often requires antibiotic selection, which limits targeted integration of multiple genes at multiple sites. To improve HDR-mediated targeted integration, while avoiding the use of selection markers, chemical treatment for increased HDR, and fluorescent enrichment of genome-edited cells was assessed in CHO cells. Chemical treatment did not improve HDR-mediated targeted integration. In contrast, fluorescent markers in Cas9 and donor constructs enable FACS enrichment, resulting in a threefold increase in the number of cells with HDR-mediated genome editing. Combined with this enrichment method, large transgenes encoding model proteins (including an antibody) were successfully targeted integrated. This approach provides a simple and fast strategy for targeted generation of stable CHO production cell lines in a rational way. Biotechnol. Bioeng. 2016;113: 2518-2523. © 2016 Wiley Periodicals, Inc.
journal_name
Biotechnol Bioengjournal_title
Biotechnology and bioengineeringauthors
Lee JS,Grav LM,Pedersen LE,Lee GM,Kildegaard HFdoi
10.1002/bit.26002subject
Has Abstractpub_date
2016-11-01 00:00:00pages
2518-23issue
11eissn
0006-3592issn
1097-0290journal_volume
113pub_type
杂志文章abstract::Electron paramagnetic resonance spectroscopy is used to observe hydroxyl radicals produced by an atmospheric pressure nonthermal plasma device at distances greater than 1 m from the discharge. The plasma device is an indirect treatment setup with closed loop airflow and hydrogen peroxide additives that is effective in...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.24853
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abstract::The human monoclonal antibody (humAb) 2F5 is a potent candidate for immunotherapy of HIV-1. The xenohybridoma derived humAb 2F5 is of IgG3 kappa isotype. Generally, IgG1 isotype has a longer half-life (beta-clearance) in humans than IgG3. Therefore the isotype was switched to an IgG1 by ligation of the 2F5 heavy chain...
journal_title:Biotechnology and bioengineering
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doi:10.1002/(sici)1097-0290(20000105)67:1<97::aid-bit1
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abstract::This work investigated the formation of microbial granules to boost the productivity of lactic acid (LA). The flocculated form of LA-producing microbial consortium, dominated by Lactobacillus sp. (91.5% of total sequence), was initially obtained in a continuous stirred-tank reactor (CSTR), which was fed with 2% glucos...
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doi:10.1002/bit.25540
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章,评审
doi:10.1002/bit.20096
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abstract::Expanded bed adsorption is a new downstream processing technique forcapture of proteins directly from unclarified feedstocks. Expanded bed adsorption reduces the number of operations in purification processes by combining clarification, concentration, and capture into one operation. It is based on stable fluidization ...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260440808
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abstract::Globally, approximately a billion patients are estimated to suffer from neurological disorders. Although there are many therapeutic candidates for the central nervous system, treatment of brain disorders is restricted by the blood-brain barrier (BBB), which is a highly selective membrane that protects the brain from e...
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
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abstract::The fim inversion system of Escherichia coli (E. coli) can behave as a unidirectional switch in an efficient manner. We have developed a new expression system for E. coli, comprising the arabinose-inducible fimE gene and the fim invertible DNA segment containing a constitutively active promoter. In this system, the ta...
journal_title:Biotechnology and bioengineering
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doi:10.1002/bit.20916
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260430209
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journal_title:Biotechnology and bioengineering
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260430505
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journal_title:Biotechnology and bioengineering
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doi:10.1002/bit.21086
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260390509
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pub_type: 杂志文章
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.10587
更新日期:2003-05-20 00:00:00
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journal_title:Biotechnology and bioengineering
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更新日期:2000-08-05 00:00:00
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
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更新日期:2017-10-01 00:00:00
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更新日期:2006-04-20 00:00:00
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journal_title:Biotechnology and bioengineering
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journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:10.1002/bit.260250310
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journal_title:Biotechnology and bioengineering
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journal_title:Biotechnology and bioengineering
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journal_title:Biotechnology and bioengineering
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journal_title:Biotechnology and bioengineering
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abstract::Bovine pancreatic trypsin was treated with ethylene glycol bis(succinic acid N-hydroxysuccinimide ester). Approximately 8 of 14 lysines per trypsin molecule were modified. This derivative (EG trypsin) was more stable than native between 30 degrees and 70 degrees C: T50 values were 59 degrees C and 46 degrees C, respec...
journal_title:Biotechnology and bioengineering
pub_type: 杂志文章
doi:
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