scholarly journals Magnetic Selection of Transiently Transfected Cells

BioTechniques ◽  
1996 ◽  
Vol 21 (5) ◽  
pp. 876-880 ◽  
Author(s):  
Stefan Schneider ◽  
Sandro Rusconi
Keyword(s):  
Transfected Cells ◽  
Selection Of

scholarly journals Enhancement of Hepatitis C Virus RNA Replication by Cell Culture-Adaptive Mutations

2001 ◽  
Vol 75 (10) ◽  
pp. 4614-4624 ◽  
Author(s):  
Nicole Krieger ◽  
Volker Lohmann ◽  
Ralf Bartenschlager
Keyword(s):  
Cell Culture ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Colony Formation ◽  
Marker Gene ◽  
Rna Replication ◽  
Clear Correlation ◽  
Transfected Cells ◽  
Adaptive Mutations ◽  
Selection Of

ABSTRACT Studies of the Hepatitis C virus (HCV) replication cycle have been made possible with the development of subgenomic selectable RNAs that replicate autonomously in cultured cells. In these replicons the region encoding the HCV structural proteins was replaced by the neomycin phosphotransferase gene, allowing the selection of transfected cells that support high-level replication of these RNAs. Subsequent analyses revealed that, within selected cells, HCV RNAs had acquired adaptive mutations that increased the efficiency of colony formation by an unknown mechanism. Using a panel of replicons that differed in their degrees of cell culture adaptation, in this study we show that adaptive mutations enhance RNA replication. Transient-transfection assays that did not require selection of transfected cells demonstrated a clear correlation between the level of adaptation and RNA replication. The highest replication level was found with an adapted replicon carrying two amino acid substitutions located in NS3 and one in NS5A that acted synergistically. In contrast, the nonadapted RNA replicated only transiently and at a low level. The correlation between the efficiency of colony formation and RNA replication was corroborated with replicons in which the selectable marker gene was replaced by the gene encoding firefly luciferase. Upon transfection of naive Huh-7 cells, the levels of luciferase activity directly reflected the replication efficiencies of the various replicon RNAs. These results show that cell culture-adaptive mutations enhance HCV RNA replication.

Magnetic Selection of Transfected Cells

2000 ◽  
pp. 210-217
Author(s):  
Gregor Siebenkotten ◽  
Ute Behrens-Jung
Keyword(s):  
Transfected Cells ◽  
Selection Of

Selection of Transfected Cells: Magnetic Affinity Cell Sorting

2003 ◽  
pp. 343-358
Author(s):  
Raji Padmanabhan ◽  
Snorri S. Thorgeirsson ◽  
R. Padmanabhan
Keyword(s):  
Cell Sorting ◽  
Transfected Cells ◽  
Selection Of

scholarly journals A simple and efficient CRISPR technique for protein tagging

2019 ◽  
Author(s):  
Fanning Zeng ◽  
Valerie Beck ◽  
Sven Schuierer ◽  
Isabelle Garnier ◽  
Carole Manneville ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Fluorescent Protein ◽  
Cell Types ◽  
Luciferase Assay ◽  
Primary Cells ◽  
Transfected Cells ◽  
Homology Directed Repair ◽  
Protein Tagging ◽  
Selection Of

Genetic knock-in using homology directed repair is an inefficient process, requiring selection of few modified cells and hindering its application to primary cells. Here we describeHomologyindependent geneTagging (HiTag), a method to tag a protein of interest by CRISPR in up to 66% transfected cells with one single electroporation. The technique has proven effective in various cell types, can be used to knock in a fluorescent protein for live cell imaging, to modify the cellular location of a target protein and to monitor levels of a protein of interest by a luciferase assay in primary cells.

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