质粒上扩增DNA的PCR条件 1、PCR reaction system 25 ng linear template （~6.5 kb） 50 pmol each primer 100 pmol each dNTP 1X Promega Taq buffer （no Mg2+） 1.5 mM MgCl2 1 U Taq DNA polymerase in 50 ul final volume 2、PCR programme 92°C / 2' 92°C / 30" 50°C or 55°C （depends on Tm of oligos） /30" 72°C / about 2' per kb Go to 2, 15 times 70°C/ 8' 4°C,hold. Takes about 2 hours to complete. 3、Notes: If you are using Pfu turbo, use its buffer （Mg already added） and decrease elongation to 1'/kb DNA. Note that this DNA is blunt ended and can be cloned directly （no purification necessary） into a phosphorylated vector. Taq made DNA needs to be AT vector cloned |
