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文献和实验Angiotensin Protein Kinase Assay
in 12 microliters kinase buffer (40 mM PIPES, pH 7.0, 10 mM MnCl2) at 4°C. 2. The kinase reaction is carried out at room temperature in a total volume of 20 microliters that contains: 15 microCi gamma-32P-ATP 5 microliters
emperically, approx 10x the concentration needed for westerns) Vortex and incubate on ice for 30 min. Meanwhile wash Protein A-Sepharose beads 2x in RIPA as follows: Cut off ends of pipetman tips Suspend beads and add beads
1 ul DNA x ul [[gamma]]-[32-P]-rATP 10 uCi T4 polynucleotide kinase 1 ul (3U/ul) 10 ul [[gamma]]-[32-P]-rATP (35020) ICN and T4 polynucleotide kinase (70031) from United States Biochemicals. 2. Incubate at 37degC
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