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:6/23/2016)... WA (PRWEB) , ... June 23, 2016 , ... ... announces the release of its second eBook, “Clinical Trials Patient Recruitment and Retention ... recruitment and retention in this eBook by providing practical tips, tools, and strategies ...
(Date:6/23/2016)... 23, 2016   Boston Biomedical , an ... designed to target cancer stemness pathways, announced that ... Orphan Drug Designation from the U.S. Food and ... cancer, including gastroesophageal junction (GEJ) cancer. Napabucasin is ... inhibit cancer stemness pathways by targeting STAT3, and ...
(Date:6/23/2016)... 23, 2016 A person commits a crime, and ... to track the criminal down. An outbreak of ... Drug Administration (FDA) uses DNA evidence to track down the ... Sound far-fetched? It,s not. The FDA has increasingly used a ... of foodborne illnesses. Put as simply as possible, whole genome ...
(Date:6/23/2016)... ... June 23, 2016 , ... ... and Mold) microbial test has received AOAC Research Institute approval 061601. , “This ... introduced last year,” stated Bob Salter, Vice President of Regulatory and Industrial Affairs. ...
Breaking Biology Technology:
(Date:5/20/2016)... -- VoiceIt is excited to announce its new marketing ... working together, VoiceIt and VoicePass will offer an ... slightly different approaches to voice biometrics, collaboration between ... Both companies ... "This marketing and technology partnership allows VoiceIt ...
(Date:5/12/2016)... DALLAS , May 12, 2016 ... has just published the overview results from the Q1 ... of the recent wave was consumers, receptivity to a ... wearables data with a health insurance company. ... choose to share," says Michael LaColla , CEO ...
(Date:5/3/2016)... , May 3, 2016  Neurotechnology, a provider ... MegaMatcher Automated Biometric Identification System (ABIS) , ... multi-biometric projects. MegaMatcher ABIS can process multiple complex ... any combination of fingerprint, face or iris biometrics. ... SDK and MegaMatcher Accelerator , which ...
Breaking Biology News(10 mins):