Carrier for Ethanol Precipitation of Nucleic Acids
Ethachinmate is a specially prepared neutral polyacrylamide polymer solution and
a useful reagent for recovering an extremely small quantity of nucleic acids using ethanol.
Wako Cat. No.
3M Sodium Acetate
0.2 mL (0.02 mL)
1.0 mL (0.1 mL)
Small Amounts of DNA/RNA Recovery
More than 20 ng/ml of DNA(>100bps) or RNA(>120bps) can be recovered.
No Incubation Process
Incubation at -20ºC or -80ºC is not necessary. It is possible to run centrifugation
immediately after the addition of Etachinmate and Ethanol, as shown in the protocol.
No Influence to Enzyme Reactions
Ethachinmate precipitates can be dissolved easily in a buffer and be used in
subsequent processes. Ethachinmate does not affect activities of DNA polymerase,
reverse transcriptase, restriction enzyme, ligase and transformation of E.coli.
When ethanol is added, Ethachinmate forms a visible pellet.
The visibility avoids the loss of extremely small amounts of nucleic acids.
DNase and RNase Free
*1) Final salt concentration should be more than 0.1 mol/L.
*2) Addition of 1 micro L of Ethachinmate per 100 micro L of DNA or RNA solution is recommended.
3 micro L of Ethachinmate is enough even if the solution amount exceeds 300 micro L. If the above process is repeated,
more addition of Ethachinmate is not necessary. (If more Ethachinmate is added repeatedly, it might increase the viscosity
of the DNA solution and affect the following process.)
*3) Vortex mixing is necessary to recover an extremely small amount of DNA or RNA.
*4) Cooling is not necessary in the centrifugation process.
*5) After centrifugation, white pellets are visible on the bottom of the tube. The precipitate can be dissolved easily in a buffer
and be used in various enzymatic reactions. The pellet can be washed 70 % ethanol if necessary.
Recovery of DNA at lower Temperature
Small concentration of DNA digested with Hind III (10 ng/ 500 micro L) was precipitated with 1 mL of ethanol
with 0.1 M sodium acetate. A dissolved precipitate was analyzed with 1 % agarose gel electrophoresis.
Lane 1 : λ/Hind III (Control)
Lane 2 : λ/Hind III + Ethachinmate 3 µL (no incubation)
Lane 3 : λ/Hind III, -20ºC, overnight.
Lane 4 : λ/Hind III, -80ºC, 20 minutes.
The results indicate that nearly 100 % of a small quantity of DNA was recovered by
the ethanol precipitation with Ethachinmate without low temperature incubation.
Recovery of Poly(A)+ RNA from Total RNA
Poly(A)+ RNA were purified from 150 micro g of total RNA extracted by ISOGEN from various mouse tissues.
The Poly(A)+ RNA was concentrated by ethanol precipitation with Ethachinmate and the concentrated
Poly(A)+ RNA was quantified by spectrophotometer at 260 nm.
||Prepared Poly(A)+ RNA
Taq DNA Polymerase
In the precipitation with Ethachinmate, 600 bps of DNA fragments were amplified by Taq DNA Polymerase
under the following conditions: 25 mM TAPS-HCL(pH 9.3), 50 mM KCI, 2 mM MgCl2, 0.2 M primers,
1 mM 2-mercaptoethanol, 0.01 % gelatin, 200 µ M dNTPs, and 1.25 uints of Taq DNA polymerase in
25 µl of reaction mixture.
The PCR was conducted 25 cycles at 94ºC for 1 min., 55ºC for 2 min and 72ºC for 1 min.
Lane 1 : Ethachinmate 0 µL
Lane 2 : Ethachinmate 0.2 µL
Lane 3 : Ethachinmate 0.5 µL
Lane 4 : Ethachinmate 1 µL
Lane 5 : Ethachinmate 3 µL
Lane 6 : Ethachinmate 5 µL
The results indicate that Ethachinmate did not inhibit any amplification
reactions of Taq DNA polymerase.