(1) Gel slice did not dissolve completely: Gel slice was too big. If using more than 300 mg of gel slice, separate it into multiple tubes. Raise temperature of incubation to 60°C and extend incubation time. (2) Incorrect DNA Elution Step: Ensure that Elution Buffer was added and absorbed to the center of the DF Column matrix (3) Incomplete DNA Elution: If the sizes of DNA fragments are larger than 10 Kb, use preheated Elution Buffer (60-70°C) during the Elution Step to improve the elution efficiency. (4) Do not overload the column with too much DNA. Higher recovery is attained when lower amount of DNA is loaded. Divide the large amount of DNA into more than one column. (5) If ddH2O is used for elution, make sure that its pH is between 7.0 and 8.5, as pH lower than 7 leads to lower elution efficiency. (6) Make sure that complete DNA elution takes place by adding no less than 30 ml of elution solution onto the membrane and letting it completely absorb into the membrane before centrifugation. (7) Large DNA fragments are eluted less readily than small DNA fragments. When the DNA product is larger than 5 Kb, use the elution solution preheated to 60ºC.
Why DNA recovery is lower than expected? "