I started to work on a special gel electrophoresis with 0.5% agarose and TPE buffer (tris-Phosphate EDTA) to visualize some encapsidated TMV rods. So I made the 10X stock solution and diluted into 1X. Both the gel and 1X solution were chilled before use. I run the gel under 150V for about 2 hrs. After the electrophoresis, the amp reading is 0.21, very high. Consequently the buffer in the tank is extremely hot. I wear double latex gloves and can barely touch the buffer. The gel becomes about 1/2 original thickness. I am very worried about the result. Wonder if anyone had any experience with TPE electrophoresis? Any good idea in reducing the heat? Maybe I can put the gel tank on ice next time? Can I switch to 0.5X TPE buffer?
Hi Tommy,
I usually work with TAE and TBE. If its low molecular weight nucleic acid that you are aiming at, then TBE is a better substitute.You can make 5X TBE stock and use 1X for running the gel. You can trying keeping Ice packs below and at the Sides of the Tank during the electrophoresis. I guess, few have even tried transferring the whole unit in the fridge and running the gel.
All the best :)
Dear Tommy,
to me, whatever buffer you are using, the heat generated is far too much and putting the unit into the fridge is asking for trouble. First, make sure you are making up the buffer right. Then make sure that your unit is not malfunctioning. TPE should have good buffering capacity so it should not overheat. make sure you are using the buffer to make up the agarose and not water and that it is the right concentration. Ensure you do not boil off your buffer, concentrating the agarose. Ensure that you are using the right voltage as the size of the tank you are using will determine what voltage should be used ie. small vs large submarine. Have a look at the link below, very infirmative:
http://cdn.intechopen.com/pdfs/35090/InTech-Discriminatory_power_of_agarose_gel_electrophoresis_in_dna_fragments_analysis.pdf
Good luck!
- Badges
- Science topic
- Similar topics
- Philosophy
- Philosophy Of Science
- Reasoning