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:2/24/2017)... (PRWEB) , ... February 24, 2017 , ... ... (TDR) procedures can be safely completed in an ambulatory surgery center (ASC) with ... (ACDF) procedures and previous two-year TDR studies. , Jake Lubinski, president of ...
(Date:2/24/2017)... ... February 24, 2017 , ... Chef Jodi Abel has returned from her ... several cities, she gained a number of delicious recipes and new techniques to share ... South Africa’s Western Cape province. It is internationally renowned for its incredible wine ...
(Date:2/24/2017)... 2017  Aethlon Medical, Inc. (Nasdaq: AEMD ... Chairman and CEO, Jim Joyce . ... last Saturday, Bill Gates warned world leaders ... than nuclear weapons. Mr. Gates expressed his concern, which ... that scientific terrorists have access to the necessary tools ...
(Date:2/24/2017)... 2017  Driven by consumers, preference towards more ... fastest growing categories, finds the recently published U.S. ... Care: Multi-regional Market Analysis and Opportunities study ... Kline. "Biotechnology actives are derived from ... effective for skin and hair care applications," explains ...
Breaking Biology Technology:
(Date:2/26/2017)... , Feb. 25, 2017  Securus Technologies, a ... solutions for public safety, investigation, corrections and monitoring, ... and Reentry. "Too often, too many ... and county jails are trying to tackle this ... and friends and family members. While significant steps are ...
(Date:2/21/2017)... LONDON , February 21, 2017 ... um 70 Millionen US-Dollar wachsen. Nach einem Gespräch mit mehr ... es einige Hindernisse zu überwinden gilt, um diese Prognose ... ... unter anderem die Mobilisierung der finanziellen Mittel für die ...
(Date:2/13/2017)... , Feb. 13, 2017 Former 9/11 Commission ... Judiciary Committee, Janice Kephart of Identity Strategy ... Donald Trump,s "Executive Order: Protecting the Nation ... 27, 2017):  "As President Trump,s ,Travel Ban, ... has now essentially banned the travel ban, it is ...
Breaking Biology News(10 mins):