题目

请问做AFLP,用EcoR1与Mse1做双酶切,酶切体系

提问时间：2021-03-26

答案

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For the combination EcoR1/Mse1,genomic DNA (75 ng) was incubated for 2 h at 37 °C with 1.25 U of Mse1,1.25 U of EcoR1,1 U of T4 DNA ligase,40 pmol of Mse1 adapters and 10 pmol EcoR1 adapters.This reaction was done in a volume of 50 µl of restrictionñligase buffer containing 10 mM Tris-HCl (pH 7.4),50 mM NaCl,1 mM dithriothreitol (DTT),0.1 mM EDTA,50 % (v/v) glycerol,0.15 % (v/v) Triton X-100 and 200 ng/µl BSA.The reaction was terminated by heating at 70 ˚C for 15 min,and then placed on ice.