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文献和实验Blunt end cloning of PCR products
End repair: Add 5-10 units of T4 DNApol and incubate at 37C for 5 minutes. Make solution to 2.5 M NH4OAc and add 1 volume of 95% ethanol; let sit for 5 minutes at RT and spin at 14K x g for 20 minutes. Wash with 70
Blunt end cloning of PCR produc
End repair: Add 5-10 units of T4 DNApol and incubate at 37C for 5 minutes. Make solution to 2.5 M NH4OAc and add 1 volume of 95% ethanol; let sit for 5 minutes at RT and spin at 14K x g for 20 minutes. Wash with 70
Blunt End Cloning of PCR Products PCR产物平端克隆
kinase. Incubate at 37ºC for 1 hour.4.Ligation reaction: Blunt end ligation is carried out in the presence of 5% PEG 8000. A microgram of Bluescript SK is blunt ended (cut with EcoRV ) and ligated with 10-100 ng of the purified PCR product in a 50μl
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