DNA extraction from filter paper - (Nov/02/2005 )
I recently received a DNA construct on a piece of filter paper. The sender dropped 2 ul of DNA onto the paper and drew a circle around the drop. I'm having trouble extracting the DNA. I was told to use 10mM Tris, ph 7.6. Does anyone have a protocol I could follow? thank you
what have you done, specifically?
I got a couple plasmids that way once. I looked up "clonesaver" online (can't remember the vendor off the top of my head). I used a variation of their protocol:
I aseptically cut out the little paper circle and stuck it in an eppy tube. I added about 30ul TE and stuffed the paper down into the buffer for good contact using the pipet tip. I vortexed it briefly, then did a quick spin to pull stuff to the bottom of the tube. I let it sit at room temp about 20-30 minutes. then, I pulled out 5ul and did a transformation of the plasmid into E coli. worked great.
OH, by the way, when done with that transformation I put the remainder in the -20 as-is. About two months later, I needed the plasmid in another strain of e coli. I pulled out the tubes, thawed them, and again transformed with 5 ul. it worked, so you can store them that way for at least a while.
How do you know that you don't have any DNA? Did you run it on a gel or try to transform it?
If you still have no luck try using a buffer with a high pH and high salt concentration (say 50mM Tris pH8.5, 1mM EDTA & 1M NaCl) then heating to 50C for 20-30 min. This should get the DNA off - this is assuming that there is any there!
DNA sequencing research
i resuspend it in 10 to 50 µl of TE and do an electroporation.