COS-7 cells are adherent to glass and plastic in culture. Their permissiveness to lytic growth of SV40 makes them a popular host for transfection with recombinant plasmids. In addition, SV40 has been a heavily researched area for many years as it has potentially been linked with some cancers, including mesothelioma, as well as being an unusually simple virus to manipulate genetically. Along with coding for its capsid proteins, SV40 contains genes for two antigens. One gene, T-antigen (or large T-antigen) is oncogenic, while the other, small t-antigen is not. T-antigen is used to control regulation and packaging of the virus when in simian cells. However, in other cell types it may interfere in cell-growth regulation, causing tumor growth.
COS-7 Cell Line Transfection Protocol
A pre-optimized COS-7 transfection kit is commercially available from Altogen Biosystems – COS-7 Transfection Kit. The kit is a lipid-based transfection reagent that is optimized to transfect miRNA, siRNA or DNA plasmid following either a standard or reverse transfection. The protocol for a 24-well plate to transfect COS-7 cells is here:
- Prepare COS-7 cell suspension:
- Detach cells using trypsin for 3-5 minutes at 37°C
- Dilute cell suspension in complete growth medium to 5 x 104 cells/mL
- Prepare transfection complexes by mixing 40 µL of serum-free medium, 5 µL of transfection reagent, and
- 600 ng DNA (or mRNA), or 30 nM – 50 nM of siRNA (or microRNA)
- Incubate transfection complexes at RT for 15 – 30 minutes
- Optional: Add 2 µL of Complex Condenser. This reagent reduces the overall size of the transfection complex, therefore increasing transfection efficiency. The Complex Condenser may increase cell toxicity
- Plate 20,000 – 30,000 cells per well in 0.5 mL of the cell suspension into culture plate
- Add prepared transfection complexes (from step 3 or 4)
- Place cells at 37ºC in a humidified CO2 incubator
- Perform the desired assay for phenotype or target gene expression 48 – 72 hours after transfection
Research Using COS-7
COS-7 is a popular research tool, and often an excellent choice for transfection experiments using recombinant plasmids. It is used to study the monkey virus SV40, as well as for other transfection experiments aimed at creating novel gene products, such as recombinant proteins. The two COS cell lines used for this type of work are COS-1 and COS-7. In order to use COS cells as a transfection host, a genetic construct preceded by the SV40 promoter is integrated into the cells. Using this strategy, the vector can be replicated by the T-antigen, which controls SV40 DNA replication and virus packaging in COS-7 cells and other simian cells.
The Simian Virus, SV40, is a popular research subject due to simplicity of the genome and ease of manipulation. Its importance also stems from association with certain types of human cancers such as mesothelioma, a cancer of the body lining. This cancer almost always develops as a result of asbestos exposure. SV40 is only 5243 nucleotides long and the DNA is arranged in a closed super helical conformation. Genetic coding for proteins of the virus’ capsid shell, as well as viral regulatory proteins, are encoded with overlapping reading frames, as well as regions coding for the T-antigen (large T-antigen) and t-antigen (small t-antigen). The T-antigen, which facilitates SV40 replication, can become bound to proteins involved in cell growth, resulting in unchecked cell growth. This unchecked cell growth can lead to tumorigenicity.
Research services – ELISA assay development using Cos-7 cells
- Replication of JC polyomavirus (JCV): JCV persists in the kidney cells post-infection and is also the causative agent of progressive multifocal leukoencephalopathy in humans. The virus itself is ubiquitous and mostly asymptomatic in humans. Scientists studied the virus’ replication in COS-7 cells, which are simian in origin. They found that it replicated efficiently in COS-7 cells, but not in COS cells without T antigen. [LINK]
- Stat5 expression and binding: After identifying the genes to encode Stat5A and Stat5B proteins in human cells, researchers were able to co-express them in COS-7 cells. They were able to identify the molecules required to activate the Stat5 proteins as well as the pathway the Stat 5 proteins followed. [LINK]
- Expression of Adipose differentiation related protein (ADRP): Researchers cloned the gene for a mouse ADRP and inserted it into a plasmid. They were able to transiently transfect the plasmid into CSO-7 cells and investigate its involvement in FA uptake. [LINK]