In transient transfection, the introduced nucleic acid exists in the cell only for a limited period of time and is not integrated into the genome. As such, transiently transfected genetic material is not passed from generation to generation during cell division, and it can be lost by environmental factors or diluted out during cell division. However, the high copy number of the transfected genetic material leads to high levels of expressed protein within the period that it exists in the cell.

Depending on the construct used, transiently expressed transgene can generally be detected for 1 to 7 days, but transiently transfected cells are typically harvested 24 to 96 hours post-transfection. Analysis of gene products may require isolation of RNA or protein for enzymatic activity assays or immunoassays. The optimal time interval depends on the cell type, research goals, and specific expression characteristics of the introduced gene, as well as the time it takes for the reporter to reach steady state. However, within a few days most of the foreign DNA is degraded by nucleases or diluted by cell division; after a week, its presence is no longer detected. Transient transfection is most efficient when supercoiled plasmid DNA is used, presumably due to its more efficient uptake by the cell. siRNAs, miRNAs, mRNAs, and even proteins can be also used for transient transfection, but as with plasmid DNA, these macromolecules need to of high quality and relatively pure (see Factors Influencing Transfection Efficiency). While transfected DNA is translocated into the nucleus for transcription, transfected RNA remains in the cytosol, where it is expressed within minutes after transfection (mRNA) or bound to mRNA to silence the expression of a target gene (siRNA and miRNA) (see Guidelines for RNA Transfection).

Transient 293 and CHO cell transfection

Large volume transient transfection of HEK293 and CHO cells adapted to suspension culture has addressed the need to obtain high amounts of recombinant protein without having to resort to the laborious process of stable cell line development.

Additional benefits of transient protein expression in mammalian cells include:

  • Rapidly go from cloned gene to large quantities of protein in 3-7 days
  • Obtain fully post-translationally modified and active mammalian proteins
  • Use your available mammalian cell culture facilities with shake flasks and a platform shaker
  • Easily purify secreted proteins from the serum free cell culture medium
  • Scale-up from 50 multi-well plates to to 25 L of culture volume and more

Featured 293 and CHO cell transfection products

Expi293™ Expression System
The Expi293™ Expression System includes Gibco® Expi293™ Expression Medium, Expi293F™ Cells, and the ExpiFectamine™ 293 Transfection Kit.  These components work together to enable high density transient transfection of Expi293F™ Cells for rapid and ultra high-yield protein production.

FreeStyle™ MAX CHO Expression System and FreeStyle™ MAX 293 Expression System
FreeStyle™ MAX Expression System combines Gibco® FreeStyle™ Media, FreeStyle™ MAX Reagent, and either FreeStyle™ CHO-S cells or FreeStyle™ 923 cells.  The FreeStyle™ MAX Systems can replace time-consuming stable cell line generation with large-scale functional protein production in one week.

FreeStyle™ MAX Reagent
FreeStyle™ MAX Transfection Reagent is a novel transfection reagent to transfection FreeStyle™ CHO and HEK-293 cells with exceptionally high efficiency to enable rapid and milligram scale of protein yield per liter.

Your partner in Transient Protein Production

Life Technologies™ supplies all required materials that are fully compatible with transient protein production including cell culture media, suspension adapted CHO and HEK-293 cells, vectors, plasmid purification products, and transfection reagents.  For researchers wishing to establish mammalian transient protein production in their laboratories, Life Technologies™ provides:

  • Starter kits, Expi293™ Expression System, FreeStyle™ MAX CHO Expression System and FreeStyle™ MAX 293 Expression System
  • Technical webinars and guidance on obtaining needed laboratory equipment
  • Technical Support
  • Hands-on training courses
  • Custom protein production services based on the above transient protein production systems