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.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Azotobacter vinelandii
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:2/11/2016)... 11, 2016 Non-profit Consortium Aims to ... to Support Research and Discovery --> ... an ambitious plan to sequence 100,000 individuals. It is intended ... at least 7 of North and East Asian countries. ... phase, the project will focus on creating phased reference genomes ...
(Date:2/11/2016)... NEW YORK , Feb. 11, 2016  Bioethics International, ... how medicines are researched, developed, marketed and made accessible to ... BMJ Open had named the publication of the ... for 2015. The publication is also featured as one of ... published in the last year that are most frequently read. ...
(Date:2/11/2016)... Feb. 11, 2016  Dovetail Genomics™ LLC today announced ... program for a planned metagenomic genome assembly service. ... metagenomic genome assembly method in a talk on Friday, ... & Technology conference in Orlando, Fla. ... complex datasets is difficult. Using its proprietary ...
(Date:2/11/2016)... ... February 11, 2016 , ... Global Stem Cells Group, ... Bankok,Thailand-based Global Stem Cells Network (GSCN) to distribute exosome injection and other biological ... Mexico, Costa Rica, Dominican Republic, Colombia, Argentina, Nicaragua, Panama, El Salvador, Venezuela, Peru, ...
Breaking Biology Technology:
(Date:2/10/2016)... India , February 10, 2016 /PRNewswire/ ... --> According to 2016 iris recognition ... identification iris recognition is more widely accepted ... available with both fingerprint and iris recognition ... the user to avoid purchasing two individual ...
(Date:2/4/2016)... , Feb. 4, 2016 The ... apparently one of the most popular hubs of ... MetaHIT and other huge studies of human microbiota, ... past few years, the microbiome space has literally ... biomedical research. This report focuses on biomedical ...
(Date:2/2/2016)... YORK , Feb. 2, 2016 /PRNewswire/ ... facilities are primarily focused on medical screening ... measure point-of-care parameters. Wearable devices that facilitate ... user,s freedom of movement are being bolstered ... for human biomedical signal acquisition coupled with ...
Breaking Biology News(10 mins):