Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation

Ruiguo Yang, Vincent Lemaître, Changjin Huang, Abbas Haddadi, Rebecca McNaughton, Horacio D. Espinosa

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Stably transfected cell lines are widely used in drug discovery and biological research to produce recombinant proteins. Generation of these cell lines requires the isolation of multiple clones, using time-consuming dilution methods, to evaluate the expression levels of the gene of interest. A new and efficient method is described for the generation of monoclonal cell lines, without the need for dilution cloning. In this new method, arrays of patterned cell colonies and single cell transfection are employed to deliver a plasmid coding for a reporter gene and conferring resistance to an antibiotic. Using a nanofountain probe electroporation system, probe positioning is achieved through a micromanipulator with sub-micron resolution and resistance-based feedback control. The array of patterned cell colonies allows for rapid selection of numerous stably transfected clonal cell lines located on the same culture well, conferring a significant advantage over slower and labor-intensive traditional methods. In addition to plasmid integration, this methodology can be seamlessly combined with CRISPR/Cas9 gene editing, paving the way for advanced cell engineering.

Original languageEnglish (US)
Article number1702495
JournalSmall
Volume14
Issue number12
DOIs
StatePublished - Mar 22 2018

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Clustered Regularly Interspaced Short Palindromic Repeats
Electroporation
Cells
Cell Line
Genes
Dilution
Plasmids
Cell engineering
Micromanipulators
Cell Engineering
Recombinant proteins
Cloning
Antibiotics
Drug Discovery
Microbial Drug Resistance
Reporter Genes
Cell culture
Recombinant Proteins
Feedback control
Transfection

Keywords

  • cell line generation
  • electroporation
  • nanofountain probe
  • single cell

ASJC Scopus subject areas

  • Biotechnology
  • Biomaterials
  • Chemistry(all)
  • Materials Science(all)
  • Engineering (miscellaneous)

Cite this

Yang, R., Lemaître, V., Huang, C., Haddadi, A., McNaughton, R., & Espinosa, H. D. (2018). Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation. Small, 14(12), [1702495]. https://doi.org/10.1002/smll.201702495

Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation. / Yang, Ruiguo; Lemaître, Vincent; Huang, Changjin; Haddadi, Abbas; McNaughton, Rebecca; Espinosa, Horacio D.

In: Small, Vol. 14, No. 12, 1702495, 22.03.2018.

Research output: Contribution to journalArticle

Yang, R, Lemaître, V, Huang, C, Haddadi, A, McNaughton, R & Espinosa, HD 2018, 'Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation', Small, vol. 14, no. 12, 1702495. https://doi.org/10.1002/smll.201702495
Yang R, Lemaître V, Huang C, Haddadi A, McNaughton R, Espinosa HD. Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation. Small. 2018 Mar 22;14(12). 1702495. https://doi.org/10.1002/smll.201702495
Yang, Ruiguo ; Lemaître, Vincent ; Huang, Changjin ; Haddadi, Abbas ; McNaughton, Rebecca ; Espinosa, Horacio D. / Monoclonal Cell Line Generation and CRISPR/Cas9 Manipulation via Single-Cell Electroporation. In: Small. 2018 ; Vol. 14, No. 12.
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