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文献和实验Preparation of BAC DNA with by Alkaline Prep Method
off supernatant and drain or wipe out excess supernatant. Should see pellet at this point. Dissolve the pellet in 700 µl of 50T/50E (50 mM Tris/50 mM EDTA, pH 8). Add 10 µl of RNase (10mg/ml) and leave at 37℃in a water bath for 1 hour. 12. Transfer the DNA
off supernatant and drain or wipe out excess supernatant. Should see pellet at this point. Dissolve the pellet in 700 µl of 50T/50E (50 mM Tris/50 mM EDTA, pH 8). Add 10 µl of RNase (10mg/ml) and leave at 37°C in a water bath for 1 hour. 11. Transfer the DNA
Protocols for ET recombination
temperature is important, usually 60℃ - 62℃ is optimal. Purify the PCR products by Qiagen column and elute with 2x 50 μl dH2O. Add 10x Dpn 1 buffer and 2 μl Dpn 1 (NEB) and digest (to eliminate template DNA) for 1 hour. Extract with Phenol:CHCl
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