Recovering DNA from Agarose Gels with Pumice

Recovering DNA from agarose gels with pumice.

Disposable device for the isolation of DNA from agarose gels.

A problem that arises when cloning DNA is the isolation of specific DNA molecules following restriction enzyme digestion or PCR amplification. Typically, a mixture of DNA fragments is electrophoretically separated by agarose gel electrophoresis, the gel is stained with ethidium bromide, and the DNA is visualized by UV transillumination. The DNA fragments of interest are excised from the gel usi...

Detection of DNA in agarose gels.

Nucleic acids that have been subjected to electrophoresis through agarose gels may be detected by staining and visualized by illumination with 300-nm UV light. Methods for staining and visualization of DNA using either ethidium bromide or SYBR Gold are described here. The most convenient and commonly used method to visualize DNA in agarose gels is staining with the fluorescent dye ethidium brom...

Quantitation of ethidium-stained closed circular DNA in agarose gels.

The fluorescence of ethidium bromide (EB) bound to equimolar amounts of supercoiled form I and unstrained linear form III pBR322, SV40 and PM2 DNA in agarose gels has been measured by scanning a photographic negative of the gel with a microdensitometer. For SV40 and PM2 DNA, commonly used staining conditions cause both forms, i.e. linear and supercoiled, to fluoresce to the same extent. This ob...

Photographic recording of fluorescent DNA bands on agarose gels.

cloning could be achieved by using this quick, alternative protocol. Not only does this Na3VO4-inhibition procedure save time, but also note that in this protocol the adjustment of vector-to-insert ratio in the ligation reaction is not necessary. To ligate with the CIAP/Na3VO4-treated vector (ca. 50 ng), we recommend using as much excess insert as is practical. Eliminating the quantitation step...