PHENOL:CHLOROFORM:ISOAMYL ALC. 25:24:1

clear yellow solution

Phenol:Chloroform:Isoamyl alcohol 25:24:1 is used in the extraction of DNA for the generation of DNA minisatellite fingerprints. It is also used for purifying RNA.

When nucleic acids are extracted with the Phenol-Chloroform-Isoamyl alcohol mixture solution, a clear component separation is achieved. The proteins present in the sample are denatured and removed (collected in the organic phase or in the interphase), while nucleic acids remain in the aqueous phase. The chloroform denatures proteins and facilitates the separation of the aqueous and organic phases, and the isoamyl alcohol reduces foaming during extraction.

Examples of the application of phenol-chloroform-isoamyl alcohol mixture are as follows:
(1) For the extraction of genomic DNA of a glycoconjugate-producing bacterium suitable for whole genome sequencing, extraction buffer and lysozyme were added to a culture of glycoconjugate-producing bacterium, which was left to stand for 10 min at 37??C, then proteinase K was added, and left to stand for 1 h at 65??C; a mixture of phenol-chloroform-isoamyl alcohol was added, and extracted; a mixture of chloroform-isoamyl alcohol was added, and extracted; cold ethanol and sodium acetate were added, and the precipitate was washed by ethanol and dried at room temperature, and RNA was added to dissolve it. centrifugation, the precipitate was washed with ethanol and dried at room temperature, and solubilized by adding DNA extraction buffer; add RNAase, and let it stand for 10 min at 37??; add PEG6000, mix well, and let it stand for 10 min at 50??; add a mixed solution of phenol-chloroform-isoamyl alcohol, and extract; add a mixed solution of chloroform-isoamyl alcohol, and extract; add a solution of cold ethanol and sodium acetate, and centrifugation, the precipitate was washed with ethanol and dried at room temperature, and the precipitate was washed with ethanol. After drying at room temperature, add ddH2O or TE buffer to dissolve, which is the extracted DNA solution.