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mixture. This mixture is then centrifuged. Because the phenol:chloroform mixture is immiscible with water, the centrifuge will cause two distinct phases to form: an upper aqueous phase, and a lower organic phase. The aqueous phase rises to the top because it is less dense than the organic phase
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The hydrophobic lipids will partition into the lower organic phase, and the proteins will remain at the interphase between the two phases, while the nucleic acids (as well as other contaminants such as salts, sugars, etc.) remain in the upper aqueous phase. The upper aqueous phase can then be
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There are some disadvantages of this technique in forensic use. It is time-consuming and uses hazardous reagents. Also, because it is a two-step process involving transfer of reagents between tubes, it is at a greater risk of
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containing the phenol:chloroform. This difference in density is why phenol, which only has a slightly higher density than water, must be mixed with chloroform to form a mixture with a much higher density than water.
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pipetted off. Care must be taken to avoid pipetting any of the organic phase or material at the
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remains in the aqueous phase. This is because DNA is more readily neutralized than RNA.
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Aqueous samples, lysed cells, or homogenised tissue are mixed with equal volumes of a
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Chan, P K S.; Chan, D. P.; To, K. F.; Yu, M. Y.; Cheung, J. L.; Cheng, A. F. (2001).
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Acid guanidinium thiocyanate-phenol-chloroform extraction
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