Home Plasmid DNA yield obtained by using Immunoport’s Mini PlusTM Plasmid DNA Extraction System is not very consistent. For the same plasmid, why do we sometimes obtain a lower yield?

There Are Several Possible Reasons Accounting For The Lower Plasmid Yield Obtained:

1) Bacterial Culture Did Not Grow Well, Thus Resulting In A Lower Cell Density. To Ensure A Well-Grown Culture, Always Inoculate Bacterial Cells From A Freshly Streaked Plate And Grow Cells In The Presence Of The Required Antibiotic(S). Ensure That Bacteria Have Grown Well After Overnight Culture And Attained An OD600 More Than 1, Meanwhile Do Not Let The Culture Grow More Than 16 Hours, Bacteria May Enter Death Phase And Plasmids In Cells Start To Be Degraded.

2) Do Not Use More Than 5 Ml Of Culture For One Preparation. Sometimes When The Culture Is Too Dense, Cells Collected From A 5 Ml Culture Cannot Be Completely Lysed. Incomplete Cell Lysis Will Lead To A Lower Yield Of Plasmid.

3) If DdH2O Of PH Less Than 7 Is Used For DNA Elution, Lower Efficiency Of Plasmid Elution Will Be Resulted.

4) When A More Concentrated Plasmid DNA Solution Is Desired, 30 Ml Of Elution Buffer Is Suggested. However, In Comparison With Using 50 Ml Elution Buffer, There Is About 40% Of Plasmid Cannot Be Eluted When 30 Ml Is Used. Therefore, No Less Than 30 MlOf Elution Solution Should Be Used.

5) Make Sure That Elution Buffer Is Absorbed Into The Membrane And To Stand The Column For 1-2 Minutes Before Centrifugation To Elute DNA. If The Buffer Still Retains On The Membrane Surface, Plasmid DNA Will Not Be Eluted Due To Lack Of Contact With Buffer. In This Case Pulse Centrifugation Of The Column For 1-2 Seconds (Do NOT Over-Centrifuge) Can Help Permeation Of The Buffer Into The Membrane.

6) Large Plasmid Is Eluted Less Readily Than Small Plasmid. When A Plasmid Is Larger Than 10 Kb, Use Elution Solution Preheated To 70˚C.