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:6/27/2016)... , June 27, 2016 /PRNewswire/ - BIOREM Inc. (TSX-V: ... been advised by its major shareholders, Clean Technology Fund ... United States based venture capital funds ... of Biorem (on a fully diluted, as converted basis), ... disposition of their entire equity holdings in Biorem to ...
(Date:6/27/2016)... ... June 27, 2016 , ... Cancer ... what they believe could be a new and helpful biomarker for malignant pleural ... Click here to read it now. , Biomarkers are components in the ...
(Date:6/27/2016)... ... June 27, 2016 , ... Newly ... technologies, services and solutions to the healthcare market. The company's primary focus is ... manufacturing, sales and marketing strategies that are necessary to help companies efficiently bring ...
(Date:6/27/2016)... 2016  Liquid Biotech USA ... a Sponsored Research Agreement with The University of ... from cancer patients.  The funding will be used ... with clinical outcomes in cancer patients undergoing a ... be employed to support the design of a ...
Breaking Biology Technology:
(Date:6/9/2016)... , June 9, 2016 ... deploy Teleste,s video security solution to ensure the safety of ... during the major tournament Teleste, an ... systems and services, announced today that its video security solution ... to back up public safety across the country. The ...
(Date:6/3/2016)... 2016 Das DOTM ... Nepal hat ein 44 Millionen ... Kennzeichen, einschließlich Personalisierung, Registrierung und IT-Infrastruktur, an ... und Implementierung von Identitätsmanagementlösungen. Zahlreiche renommierte internationale ... teilgenommen, aber Decatur wurde als konformste und ...
(Date:6/2/2016)... 2016   The Weather Company , an IBM Business ... industry-first capability in which consumers will be able to interact ... questions via voice or text and receive relevant information about ... Marketers have long sought an advertising solution that can ... personal, relevant and valuable; and can scale across millions of ...
Breaking Biology News(10 mins):