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AlanFung:LabNotes/Human MDA Halotyping/2009-7-20
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==Procedure== *Harvest the cells and transfer them to a 1.5ml tube *Centrifuge at 16000g for 10sec to pellet the cells *Remove the supernatant *Add 200ul pbs to wash the cells repeat centrifuge and remove pbs *Vortex vigorously to resuspend cells *Add 600 ul of nuclei lysis solution and pipet to lyse the cells (pipet until no visible cell clumps remain) *add 3ul of rnase solution to the nuclear lysate and mix the sample by inverting the tube 5 times uncubate the mixture for 30m 37C allow the sample to cool to rt for 5m before proceeding *add 200ul of protein precipitation solution and vertex vugorously at high speed for 20sec *Chill sample on ice for 5m *centrifuge for 4n at 16000g *add 600ul rt isopropanol to a 1.5ml tube *transfer the supernatant containing the DNA to the 1.5ml tube *gently mix the solution by inversion until the white thread like strands of dna form a visible mass *centrifuge for 1m at 16000g at rt *decant the supertant *add 600ul of rt 70% ethanol and gently invert the tube several times to wash the dna *centrifuge for 1m at 16000g at rt *decant the supernatant *invert the tube on clean absorbent paper and air dry the pellet for 15m *add 100ul of dna rehydration solution *incubate at 65C for 1 hr periodialy mix the solution by gently tapping the tube *store dna at 4C *nanodrop
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