Phenol – chloroform – isoamyl alcohol mixture

Phenol-chloroform-isoamyl alcohol mixture (PCI-mixture) is a widely utilized mixture in molecular biology and biochemical research, specifically designed for nucleic acid purification, including DNA and RNA. With its essential composition of 25% phenol, 24% chloroform, and 1% isoamyl alcohol, PCI-mixture plays a pivotal role in separating proteins from nucleic acids during the purification process. Phenol denatures proteins, chloroform creates a distinct layer to trap them, and isoamyl alcohol facilitates the clear separation of these layers, enabling the isolation of purified DNA or RNA in the aqueous phase.
The applications of PCI-mixture are diverse, encompassing RNA isolation, where it efficiently extracts RNA from animal and plant tissues for downstream analyses like reverse transcription and gene expression studies. Further, PCI-mixture also proves instrumental in DNA extraction, enabling efficient recovery for various applications such as DNA minisatellite fingerprinting for genetic identification. As a general purification technique, PCI-mixture ensures high-quality preparations of both DNA and RNA for further research.

Phenol − chloroform − isoamyl alcohol mixture has been used in nucleic acid extraction in bile duct tissue[1], Drosophila[2], chromatin and RNA immunoprecipitation in Caenorhabditis elegans[3] and mitochondrial DNA.[4]Further, also in the extraction of DNA for the generation of DNA minisatellite fingerprints[5]

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.

mixture of phenol; chloroform; isoamyl alcohol in the ratio of 25:24:1 (v/v/v); saturated with 100 mM TRIS pH 8.0; contains ~0.1% 8-hydroxyquinoline

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