Navigation Links
Azotobacter vinelandii

Multiporator / Electroporator 2510 Transformation Protocol

Protocol No. 4308 915.503 04/2002

Microorganism Azotobacter vinelandii Cell type Bacteria, gram negative Molecules injected Plasmid DNA Growth medium Nitrogen-free medium Washing solution Ice-cold 10% sterile glycerol Electroporation solution Ice-cold 10% sterile glycerol Outgrowth medium Azotobacter growth (AG) medium Cuvette 1 mm gap width Reference Kornyi, P. et al 1998 Research in Microbiology 149 361-372

Making electrocompetent cells:

  1. Cultivate cells with vigorous shaking at 30 C to an O.D.620 of 0.4-0.5.

  2. Harvest by centrifugation at 5,000 rpm for 10 min at 0 C.

  3. Wash with the original culture volume of ice-cold 10% glycerol.

  4. Repeat this step three times using a half and a quarter of the original volume, and finally 2-4 ml of the glycerol solution (7.0 x 107 cells/ml final cell concentration).

  5. Freeze 40 l aliquots in liquid nitrogen and store at 70 C.

Electroporation of cells:

  1. Add 1 l (300 ng/l) plasmid DNA to 40 l of electrocompetent cells. Homogenize by gently mixing with pipette

  2. several times. Transfer mixture into a prechilled cuvette.
  3. Wipe moisture from the cuvette and insert the cuvette into the device.

  4. Electroporation:

  5. Mode Prokaryotes O Voltage (V) 1,800 V Time constant (T)
    5 ms No. of pulses (n) 1
  6. Immediately add 1 ml AG medium and incubate with shaking at 200 rpm for 1 h at 30 C.

  7. Plate onto selective AG media plates.

Expected results:

Transformation efficiency up to 6.8 x 105 transformants/g of DNA.



Page: All 1 2

Related biology technology :

1. Azotobacter vinelandii
Post Your Comments:

(Date:11/27/2015)... ... November 27, 2015 , ... Pittcon ... over 2,000 technical presentations offered in symposia, oral sessions, workshops, awards, and posters. ... a wide range of applications such as, but not limited to, biotechnology, biomedical, ...
(Date:11/25/2015)... --> ... 2020 report analyzes that automating biobanking workflow will ... long-term samples, minimizing manual errors, improving the workflow ... errors such as mislabeling or inaccurate sample barcoding ... a vital role in blood fractionation, DNA extraction, ...
(Date:11/25/2015)... and HOLLISTON, Mass. , ... Inc. (Nasdaq: HART ), a biotechnology company developing ... CEO Jim McGorry will present at the ... 1, 2015 at 2:30 p.m. PT. The presentation will ... for 30 days. Management will also be available at ...
(Date:11/25/2015)... , Nov. 25, 2015 Orexigen® Therapeutics, ... will participate in a fireside chat discussion at the ... New York . The discussion is scheduled for ... .  A replay will be available ... Contact:McDavid Stilwell  , Julie NormartVP, Corporate Communications and ...
Breaking Biology Technology:
(Date:11/12/2015)... , Nov. 11, 2015   Growing ... reliable analytical tools has been paving the way ... qualitative determination of discrete analytes in clinical, agricultural, ... are being predominantly used in medical applications, however, ... environmental sectors due to continuous emphasis on improving ...
(Date:11/9/2015)... Nov. 9, 2015  Synaptics Inc. (NASDAQ: SYNA ... announced broader entry into the automotive market with a ... the pace of consumer electronics human interface innovation. Synaptics, ... ideal for the automotive industry and will be implemented ... Europe , Japan , ...
(Date:10/29/2015)... Daon, a global leader in mobile biometric ... new version of its IdentityX Platform , IdentityX ... have already installed IdentityX v4.0 and are ... FIDO UAF certified server component as an option ... features. These customers include some of the largest and ...
Breaking Biology News(10 mins):