Hi everyone,
I just wonder that I combined the DNA mixture with Lipofectamine in the Eppendorf tube, then I stand it at room temperature for 20-30 min (as protocol) in the clean bench of cell culture room. However, I am crazy that I turn on the UV light and let them exposed for 20min.
I wonder that the UV light can penetrate to the Eppendorf tube and affect to my DNA mixture (because I may use it for transfection)?
Thank you.
Hello Nuoc Non Tran
Eppendorf tubes are not UV safe as they cannot block UV light. These tubes need to be protected from sunlight and UV rays.
Cell culture rooms are equipped with germicidal UV lamps that radiate light at a wavelength of 254nm (UV-C band) for decontamination as it destroys harmful micro-organisms by attacking their DNA. UV-C is the most damaging type of UV radiation.
Both long and short wavelength UV light are damaging to DNA, but in different ways. Short wavelength UV-B and UV-C can directly cause dimerization of pyrimidines, and directly prevent replication of plasmid DNA. Long wavelength UV-A is less damaging and causes mutations through the production of reactive oxygen species. UV-A is less harmful to naked DNA. However, with enough exposure, UV-A could also damage DNA.
Twenty minutes of UV exposure could be enough to damage the DNA if the UV lamp is effective and working well. However, since the UV light is in the culture room, the distance of the tube from the UV source will also make a difference as the intensity of UV radiation will be reduced. So, you can just give it a try. Maybe it might work!
Good Luck.
Malcolm Nobre Thank you very much for your detail information. It is really helpful for me. I already harvested the cell lysate, let see the result. In parallel, I also made a new set of samples. ^_^
Don't panic. Polyethylene is absorbing pretty well below 300nm (see e.g.Article Polymer nanocomposites part 2: Optimization of zinc oxide/hi...
). If some damage has occurred, it is not uniform, so your transfection should be ok. You might want to be more concerned about potential damage to your skin and eyes, if you have been exposed. Did you get sunburn?
According to the manufacturer (https://www.eppendorf.com/product-media/doc/en/086008_Operating-Manual/Eppendorf_Consumables_Instructions-use_Tubes.pdf) Eppendorf tubes are made from coloured or colourless polypropylene (PP).
The UV lamp in the cabinet you mention, is most likely a germicidal UV-C lamp, and PP is known to be only partially absorbent of UV-C radiation in the wavelengths (240 - 270 nm) that will cause damage to DNA.
Therefore, thin-walled PP Eppendorf tubes may not fully block UV-C radiation, as can be seen in the comparative transmission spectra shown here (for PMMA, PP, PC and PET): https://ars.els-cdn.com/content/image/1-s2.0-S0048969720326437-gr1_lrg.jpg in this publication:Article Material selection and prediction of solar irradiance in pla...
Therefore, 20 min of UV-C irradiation of these small Eppendorf tubes, at close range in a cabinet, is likely to have caused some significant deterioration (ie: dimerisation and crosslinking of the DNA sample). However, as already mentioned by others, the sample is not likely to be uniformly damaged and therefore a significant proportion of your DNA sample may still be suitable for your transfection application.
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