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SafeView can be used as a replacement for ethidium bromide as they both work on general agarose. We recommend using SafeGreen for downstream cloning applications as SafeView can interfere with the ligation reaction, yielding fewer colonies.
There are fluorescent compounds in SafeView and these fluorescent compounds have the capability to bind to DNA. There may be some unknown effects of SafeView that have not been documented but that applies to the SYBR set as well; however, SafeView products are definitely not as carcinogenic as ethidium bromide.
The Safe-(Red, Green and White) loading dyes work the same as 6x loading dye, loaded with the sample. SafeView Classic is used directly in the gel and the running buffer.
Under UV, SafeView Classic emits a green fluorescence when bound to both single and double stranded DNA templates, therefore they cannot be differentiated by this method. It will emit a red fluorescence when bound to RNA templates.
The SafeView Stains (Red, Green and White) do not perform in this way and will stain all nucleic acid templates one color.
All the SafeView products should be stored at four degrees Celsius.
Under UV light, SafeView Classic emits a green fluroescence when bound to both single and double stranded DNA templates. It will emit a red fluorescence when bound to RNA templates. No filter is necessary for viewing these colours, however a filter may be needed for photographing the gel.
Our in-house testing has shown that SafeView stained gels (>10ng DNA loaded per lane) can still be effectively visualized up to 1 week later with only a slight decrease in brightness. Gels should be stored properly to maintain a good signal, at 4C in the dark, sealed in a plastic bag or pouch with wet paper towel loosely wrapped around the gel.
Make sure you are following the protocol carefully and adding 5ul of SafeView for every 100ml of running buffer and gel. It is critical that both buffer and gel have SafeView dye in them otherwise it will not work. This is different from ethidium bromide.
It will take one hour of washing with water.
2 hours over 100C
Safe-Green has Excitation Wavelength of 290nm and Emission Wavelength of 490nm, and its sensitivity is range between 0.2-0.6ng
Safe-Red has Excitation Wavelength of 490nm and Emission Wavelength of 630nm, and its sensitivity is range between 0.3-0.8ng
Safe-White has Excitation Wavelength of 320nm and Emission Wavelength of 480nm, and its sensitivity is range between 0.2-0.5ng
SafeView Classic emits green fluorescence when bound to dsDNA and ssDNA and red fluorescence when bound to RNA. This stain has one excitation (290 nm) and two emission spectra (490 nm and 605 nm) and the sensitivity range is between 0.1-0.3ng.
SafeView Plus has Excitation Wavelength of 490nm and Emission Wavelength of 520nm and the its sensitivity is range between 0.05-0.1ng
Only SafeView Plus (G468) should be used in a post stain. SafeView classic and Safe Stains are not designed for post-staining. SafeView Classic must be added to the gel and the running buffer prior to the loading of the samples. Safe-(Red, Green and White) stains must be added to the sample before loading it to the gel.
A reason for this is that most gel doc systems have been optimized for EtBr so that is why the EtBr signal is so strong in the pictures.
Shifting is unavoidable and quite natural for such fragments, regardless of the staining any . We suggest to use SafeGreen ladders, which will give accurate molecular weight with no additional staining agents needed.
It is very difficult to detect 100bp and 200bp bands in 1% gel with any stains. Higher gel concentrations should be used, such as 2% agarose.
Yes, we have tested our SafeView products for this application.
Note: All prices in US Dollars