α4-200™ plasmid purification kit
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  • α4-200™ plasmid purification kit

α4-200™ plasmid purification kit

$85.00
Delivered within 1-3 days

Catalog: AUK1.

Equivalent of 200 mini-preps.

Our unique kit is designed for purification of up to 35 k.b. plasmid DNA from E. coli cells. 

It utilizes a different approach than other commercial kits do. 

The entire process of plasmid purification by this kit happens in a single tube and involves fewer steps than micro-column-utilizing kits.  Consequently, our kit assures lower per-sample cost of purified DNA and demands less operator involvement time while producing DNA suitable for all types of enzymatic treatments and efficient transformation of all types of cells, including mammalian. Additionally, the kit allows easy up-scaling of the procedure depending on the amount of DNA required.

Quantity

a4-200™ plasmid purification kit.

Protocol

Prior to using the kit for the first time, add 9 mL of 96% ethanol to the bottle labeled “Solution #3”!!!

Store the kit in the refrigerator.

  1. Collect cells from overnight E. coli cell cultures in microcentrifuge tubes provided with the kit by centrifugation (two (2) minutes at 13 rsf).
  2. Remove the majority of supernatant, leaving 20 – 30 mL for further resuspension of cells. We recommend decanting the majority of the supernatant to waste and adding 1 – 2 vigorous shakes into the waste receptacle.

Note: Volumes listed below should be used for cells collected from 1.5 – 3.0 mL of cell culture. One provided tube may be used to purify plasmid DNA from up to 15 mL of cell culture, however, volumes of used solutions should be increased proportionally.

  1. Transfer tubes to 80 °C (or 100 °C) and incubate for 15 minutes (or 2 minutes in case of 100 °C).
  2. Re-suspend cells in the remaining (~30 µl) supernatant using a Vortex.
  3. Add 100 µl of Solution #1 to each tube.
  4. Thoroughly mix the contents of the tubes (a short burst on the Vortex should suffice), then incubate the tubes at 37 °C for 5 – 15 minutes.
  5. Centrifuge the tubes for 10 – 20 minutes at 13 rsf to achieve compaction of the solid fraction to about 1/3 – 1/5 of the total volume.
  6. Remove pellets from tubes by inserting a sterile toothpick (provided in the kit) into each pellet, spinning it inside of pellet, and pulling the toothpick out of the tube together with the stuck pellet. Note: We recommend using flame-sterilized metal forceps to remove toothpicks from the storage container.
  7. Add 150 µl of Solution #2 to the remaining supernatant of each tube and mix the contents by gently inverting the tubes several times (for plasmids larger than 20 k.b.) or by a short burst on a Vortex.
  8. Centrifuge tubes for 5 minutes at 13 rsf.
  9. Remove supernatant by aspiration.
    Note: We recommend using a faucet aspirator/vacuum pump connected to a 10-    mL pipet tip. Move the tip along the side opposite where the pellet should be located. We found that the same tip can be used for multiple samples if briefly washed with fresh deionized water.
  10. Add 50 µl of Solution #3 (which was previously diluted and mixed with ethanol) to each tube.
  11. Centrifuge tubes for 10 seconds at 13 rsf.
  12. Remove supernatant by aspiration.
  13. Repeat steps 12 – 14 one more time.
  14. Add 50 µl of Solution #4 to each tube and incubate tubes at 37 °C for 5 – 10 minutes.
  15. The purified DNA is ready for further manipulations!

AUK1
200 Items