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The cloning mixture was desalted by dialyzing against nuclease-free water using a 0.05 m membrane filter (Merck, VMWP02500) for 30 minutes

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In addition, P 2 was extracted with three times volume of PBS (0.1 mol/L, pH 7.0, containing 1.0 mol/L NaCl), at 4C for 18h and the solution was continuously stirred
