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Published TCIMAIL newest issue No.200
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With the improvement of analytical techniques at the single molecule and single cell level, and because RNA is sometimes a target for nucleic acid drugs, RNA imaging has become a very important tool in molecular biology and drug discovery research. Here we introduce products effective for RNA imaging.
Advantages
- Stains RNA in bacteria.
- Wavelengths of RNA stained with T-Fluor 480: λex, max = 450 nm, λem, max = 485 nm
- Very high fluorescence intensity. The signal can be observed without washing.
- Tested for RNA staining.
- T-Fluor 480 Solution (Product No. T4436) is filter-sterilized and convenient to use in liquid form.
Application: RNA staining of E. coli with T-Fluor 480
- Cultivate E. coli BW25113 on LB medium at 37°C overnight.
- Add 1/100 volume of culture medium to LB medium with or without 100 µg/mL rifampicin (*) and incubate at 37°C for 4 hours.
- After removing the supernatant by centrifugation, add PBS containing 6.5 μg/mL T-Fluor 480 to the pellet and stain at room temperature for 5 minutes.
- Observed under a fluorescence microscope using a filter for GFP.
* Rifampicin: RNA synthesis inhibitor
RNA staining was observed in rifampicin(-) medium where RNA synthesis was not inhibited.
(The images were provided by Dr. Shinya Sugimoto.)
Application: Observation of RNA in transparent biofilm
- Incubate Methicillin-resistant Staphylococcus aureus (MRSA) in a glass-bottomed dish for 24 hours to form a biofilm.
- Remove the culture medium and suspended bacteria. Then fix and wash the biofilm.
- Add 8 µg/mL T-Fluor 480 solution in PBS to the dish, and stain the biofilm at room temperature for 30 minutes.
- Add PBS or biofilm clearing reagent iCBiofilm-H1 (Product No. T4031) to the dish, and observe the biofilm.
The addition of iCBiofilm-H1 made the biofilm transparent, and RNA deep in the biofilm was observed.
(The images were provided by Dr. Shinya Sugimoto.)
Fluorescence spectra
Fluorescence spectra of T-Fluor 480 without reaction of RNA
Fluorescence spectra of T-Fluor 480 after reaction of RNA
