advertisement

You do not have JavaScript enabled or are using an older version of Adobe Flash. You can update your version of Adobe Flash here.

Nocardia Asteroides

ShareThis

Nocardia asteroides
Multiporator / Electroporator 2510
Transformation Protocol Protocol No. 4308 915.525 – 01/2002
Microorganism Nocardia asteroides
Cell type Bacteria, gram positive
Molecules injected Plasmid DNA (in TE buffer)
Growth medium N2 medium (20 g/l glucose, 2 g/l yeast extract, 4 g/l beef extract, 6 g/l tryptone, 2 g/l NaCl, 10 g/l glycine)
Washing solution Distilled water
Electroporation solution 0.3 M sucrose, 15% glycerol
Outgrowth medium N2 medium (without antibiotics)
Cuvette 2 mm gap width
Reference Yao W., et al • 1994 • Current Microbiology 29 • 223-227
Making electrocompetent cells:

1. Inoculate a fresh overnight culture of bacteria into N2 medium. Grow cells at 28 °C with shaking to an O.D.600 of 0.7.
2. Harvest by centrifugation.
3. Wash twice with _ volume of distilled water.
4. Resuspend in a final volume of 1/20 of the original volume of electroporation solution.

Electroporation of cells:

  1. Add 2 µl (0.1 µg) plasmid DNA to 50 µl of electrocompetent cells. Homogenize by gently mixing with pipette several times. Transfer mixture into a prechilled cuvette.
  2. Wipe moisture from the cuvette and insert the cuvette into the device.
  3. Electroporation:

    Mode Prokaryotes “O“
    Voltage (V) 2,500 V
    Time constant (T) 5 ms

  4. Add 1 ml N2 medium and incubate for 2 hours at 30 °C.
  5. Plate cells on selective YEME medium with a 3 ml YEME soft agar (0.4%) overlayment.
Expected Results:
Transformation efficiency up to 8 x 104 transformants/µg of DNA.

Contact Information

In the United States:
Eppendorf North America, Inc.
102 Motor Parkway,
Hauppauge, NY 11788-5178
Tel: 800-645-3050
Fax: 516-334-7506
Web Site: http://www.eppendorfna.com/

Outside the United States:
Eppendorf AG
Barkhausenweg 1
22339 Hamburg
Germany

Customer Service: ++ 49 40 53 801-0

Fax Number: ++ 49 40 53 801-556

Article Tools
  • Email a Colleague
  • Print
  • ShareThis
advertisement

You do not have JavaScript enabled or are using an older version of Adobe Flash. You can update your version of Adobe Flash here.

Email Newsletter Sign-Up

Stay updated on the latest technologies and news with Biocompare's newsletters
(See samples here)












 Select All

Privacy Policy | Manage Subscriptions

Loading

Loading