Navigation Links
Staphylococcus aureus

Multiporator / Electroporator 2510 Transformation Protocol Protocol No. 4308 915.533 01/2002 Microorganism Staphylococcus aureus Cell type Bacteria, gram positive Molecules injected Plasmid DNA (pC194) Growth medium Basic medium (BM: 1% peptone, 0.5% yeast extract, 0.1% glucose, 0.5% NaCl,
0.1% K2HPO4) Washing solution 10% glycerol (in distilled water) Electroporation solution 10% glycerol (in distilled water) Outgrowth medium SMMP50 medium (5.5 parts SMM buffer (1 M sucrose, 0.04 M maleic acid, 0.04 M MgCl2, pH 6.5), 4 parts 7% Panassy broth, 0.5 parts 10% BSA) Cuvette 2 mm gap width Reference Augustin J. and Gtz F. 1990 FEMS Microbiology Letters 66 203-208 Making electrocompetent cells:

1. Inoculate a flask of BM with a fresh overnight culture. Grow at 37 C with shaking to an O.D.578 of about 0.5 to 0.65. 2. Wash in one volume of 10% glycerol at room temperature, followed by washes with 1/2, 1/20 and 1/50 vol umes. 3. Resuspend to final volume of 1-5 x 1010 cells/ml in 10% glycerol. Aliquote and freeze at 70 C.

Electroporation of cells:

  1. Thaw cells for 5 min at room temperature. Add 1-2 l plasmid DNA (100-500 ng) to 50 l of electrocompetent cells. Homogenize by gently mixing with pipette several times. Incubate at room temperature for 30 minutes.
  2. Transfer mixture into a cuvette and insert the cuvette into the device.
  3. Electroporation:

    Mode Prokaryotes O Voltage (V) 1,800 V Time constant (T) 5 ms
  4. Add 1 ml SMMP50 medium to the cells and transfer to sterile tube. Incubate 90 minutes at 37 C with moderate shaking.
  5. Plate on selective BM plates. Incubate 20 hours at 37 C.
Expected Results: Transformation efficiency up to 1.5 x 105 transformants/g of DNA.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Staphylococcus epidermidis
2. Staphylococcus aureus
3. Staphylococcus epidermidis
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:6/27/2016)... June 27, 2016  Global demand for enzymes ... through 2020 to $7.2 billion.  This market includes ... cleaning products, biofuel production, animal feed, and other ... and biocatalysts). Food and beverages will remain the ... increasing consumption of products containing enzymes in developing ...
(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 , ... Rolf K. ... the faculty of the University of North Carolina Kenan-Flagler Business School ... entrepreneurship at UNC Kenan-Flagler, with a focus on the school’s international efforts, leading ...
(Date:6/24/2016)... discussions on a range of subjects including policies, debt and ... Poloz. Speaking at a lecture to the Canadian ... to the country,s inflation target, which is set by both ... "In certain areas there needs to be ... why not sit down and address strategy together?" ...
Breaking Biology Technology:
(Date:4/26/2016)... LONDON , April 26, 2016 ... a product subsidiary of Infosys (NYSE: ... to integrate the Onegini mobile security platform with ... http://photos.prnewswire.com/prnh/20151104/283829LOGO ) The integration will ... to access and transact across channels. Using this ...
(Date:4/15/2016)... 15, 2016  A new partnership announced today ... underwriting decisions in a fraction of the time ... and high-value life insurance policies to consumers without ... With Force Diagnostics, rapid testing (A1C, Cotinine and ... (blood pressure, weight, pulse, BMI, and activity data) ...
(Date:4/13/2016)... -- IMPOWER physicians supporting Medicaid patients in Central ... in telehealth thanks to a new partnership with higi. ... patients can routinely track key health measurements, such as ... when they opt in, share them with IMPOWER clinicians ... retail location at no cost. By leveraging this data, ...
Breaking Biology News(10 mins):