Genomic DNA extraction method.
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Last Update: 2020-10-21
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Source: Internet
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Author: User
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1, wall cells digested with trypsin, centrifugal collection.
2, cells re-suspended in the cold PBS rinsing once, centrifugal collection.
3, repeat 2.
4, add 5ml
. DNA
a buffer (10m mol/L Tris-cl, 0.1 mol/L
EDTA
,0.5%
SDS
), mixed well.
5, add 25ul
protease
K, so that the final concentration of 100ug/ml, mixed well, 50 degrees C water bath 3h.
6, with the same volume of phenol pumping once, 2500r/min centrifugation to collect water phase. The water phase is collected at 2500r/min centrifugation with an equal volume (phenol, chloroform, isosterol) mixture.
is used in the same volume of chloroform, isosterol pumped once.
7, add the same volume of 5mol/L LiCL, mix well, ice bath, 10min.
8, 2500r/min, centrifugal 10min. Turned into a
centrifugal tube
tube. Add isopropyl alcohol of equal volume. Room temperature for 10 minutes.
9, 2500r/min, centrifuged 10min. Discard the Qing. Add 0.1x volume 3mol/L sodium acetate (PH5.2) and 2x volume-20C pre-cooled waterless ethanol. -20 degrees C for 20 minutes.
10, 12000r/min, centrifuged at room temperature for 5 minutes. Discard the Qing. Dissolve the DNA in an appropriate amount of TE.
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