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Application notes
Himac Application No. 59 – Separation of Plasmid DNA


Himac Preparative Micro-Ultracentrifuge CS Series Neo-Angle Rotor S120NT
We've shown in our himac APPLICATION it takes 3h to separate plasmid DNA by S120NT neo-angle rotor (holding 2ml tube)1). Here we tried this separation even shorter.Then we can prove this separation for 2 hours.


Results



Conditions

(1)   Condition of ultracentrifugation


Rotor

Speed (rpm)

Time(h)

Temp.(?C)

     Acc. mode

Dec. mode

S120NT

neo-angle rotor

120,000

1.5

1.75

2.0

20

"9"


"7"



(2)   Used tube : 2PA Cone-Top tube*
*Registered trade mark of Seton Scientific Company.

(3)   Sample preparation
We used E.coli., JM109 holding plasmid pUC19 DNA. The E.coli., JM109 was cultured over night and crude DNA was isolated by alkaline-SDS method and other processes and dissolved with TE buffer. We used this solution as a sample.


For each 2PA Cone-Top tube*, we provided
Sample: 1.48ml
CsCl   : 1.52g
Ethydium Bromide (10mg/ml) : 50 μl
Polyethylen (10) octylphenylether (Triton X - 100) : 1 - 2 μl
We mixed these and poured the resulting mix into a tube. If this is not enough to fill the tube, the tube should be filled with an additional filler solution prepared in advance (that dissolve 0.93g of CsCl to 1ml of TE buffer).

Reference
1) himac APPLICATION No.46 (1995).

Product information Himac Ultracentrifuge:



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