[생물학실험] QIAGEN Midi, Maxi prep 전처리 과정

Step2, Step3에 관한 QIAGEN protocol

Procedure

1. Step2. Mini culture (Midi, Maxi 공통 전처리 과정) > Pick a single colony from a freshly streaked selective plate and inoculate a starter culture of 2–5 ml LB medium containing the appropriate selective antibiotic. Incubate for approx. 8 h at 37°C with vigorous shaking (approx. 300 rpm).
2. Use a tube or flask with a volume of at least 4 times the volume of the culture.
3. Step3. Dilute the starter culture 1/500 to 1/1000 into selective LB medium.
   • Midi bacterial culture > For high-copy plasmids, inoculate 25 ml medium with 25-50ul of starter culture, For low-copy plasmids, inoculate 100 ml medium with 100-200ul of starter culture.
   • Maxi bacterial culture > For high-copy plasmids, inoculate 100 ml medium with 100-200ul of starter culture, For low-copy plasmids, inoculate 500 ml medium with 250-500ul of starter culture.
   Grow at 37°C for 12–16 h with vigorous shaking (approx. 300 rpm).
4. Use a flask or vessel with a volume of at least 4 times the volume of the culture. The culture should reach a cell density of approximately 3–4 x 109 cells per milliliter, which typically corresponds to a pellet wet weight of approximately 3 g/liter medium.
5. Harvest the bacterial cells by centrifugation at 6000 x g for 15 min at 4°C.
6. If you wish to stop the protocol and continue later, freeze the cell pellets at –20°C

QIAGEN-Plasmid-Purification-Handbook--Miniculture-Midiculture_April-2012.pdf

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