After overnight (12-16 hrs) incubation of E. coli in LB, when I attempted to extract DNA using Kit, it yielded a low concentration of DNA (10-20 ng/µL). Can anyone help me out to improve its concentration (>100ng/uL).
Kits normally give a much concentration than what you got.
Things to check:
- E. coli was grown using appropriate antibiotic selection for the plasmid
- kit components are not expired
- volume of cell culture within parameters of kit
- cells were entirely lysed (some kits have a blue color indicator)
Hope one or more of those will help!
What size is your plasmid? Also what volume of culture you using? A common cause for low yields is cracked zeolite resin in the columns. This can occur with kit age or if you jab it using a pipette tip. Other tips for increasing yield are to put your lysate and elutant though the column twice and to have a 2 min incubation when eluting.
How did the cell cultures look after the incubation? Nice and cloudy? If there are few cells going in then you'll get a low yield. Is anyone else having the same problem using the same kit? As mentioned above, kits do expire. It can help to warm the elution buffer (50C) then let it incubate for a minute on your column prior to eluting.
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