Navigation Links
Borrelia burgdorferi

Multiporator / Electroporator 2510 Transformation Protocol Protocol No. 4308 915.506 12/2001 Microorganism Borrelia burgdorferi B31 Cell type Bacteria, gram negative Molecules injected Linear DNA (in water) Growth medium Barbour-Stoenner-Kelly (BSK) II medium without gelatin Washing solution PBS, without divalent cations; EPS (272 mM sucrose, 15% glycerol) Electroporation solution EPS (272 mM sucrose, 15% glycerol) Outgrowth medium BSK II medium Cuvette 2 mm gap width Reference Samuels D.S., et al 1994 Journal of Bacteriology 176 6045-6049

Making electrocompetent cells:

  1. Grow cells in 500 ml BSK II at 32 C to a cell density of 3-7 x 107 cells/ml.

  2. Harvest by centrifugation (4,000 x g, 20 min., 4 C). Wash twice in 60 ml cold PBS and pellet by centrifugation (3,000 x g, 10 min., 4 C).

  3. Wash three times in 20 ml cold EPS and pellet by centrifugation (2,000 x g, 10 min., 4 C).

  4. Resuspend the cells in 0.6 ml cold EPS.

Electroporation of c ells:

  1. Add 0.3 to 1 g DNA to 50 l of electrocompetent cells on ice. 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:

  4. Mode Prokaryotes O Voltage (V) 2,500 V Time constant (T) 5 ms
  5. Immediately add 1 ml BSK II medium, transfer cells to a 15 ml tube and add another 9 ml of BSK II. Incubate for 24-48 hours at 32 C with shaking.
  6. Plate 0.1 ml of culture on solid BSK II medium. Pellet the remaining 9.9 ml (4,000 x g, 10 min.), resuspend in 1 ml of supernatant and plate on selective plates. Incubate for 14 days, at 32 C in a humidified 5% CO2 chamber.

Expected results:

Transformation efficiency up to 103 transformants/g of DNA.



Page: All 1 2

Related biology technology :

1. Borrelia burgdorferi
Post Your Comments:

(Date:11/25/2015)... , Nov. 25, 2015 /PRNewswire/ - Aeterna Zentaris Inc. ... its business and prospects remain fundamentally strong and ... (zoptarelin doxorubicin) recently received DSMB recommendation to continue ... following review of the final interim efficacy and ... Primary Endpoint in men with heavily pretreated castration- ...
(Date:11/25/2015)... ... November 25, 2015 , ... Jessica Richman and Zachary Apte, ... their initial angel funding process. Now, they are paying it forward to other ... stage investments in the microbiome space. In this, they join other successful ...
(Date:11/24/2015)... SAN DIEGO , Nov. 24, 2015 Halozyme Therapeutics, ... Jaffray Healthcare Conference in New York on ... Dr. Helen Torley , president and CEO, will provide a ... New York at 1:00 p.m. ET/10:00 a.m. ... communication and investor relations, will provide a corporate overview. --> ...
(Date:11/24/2015)... 24, 2015 /PRNewswire/ - Aeterna Zentaris Inc. (NASDAQ:  AEZS) (TSX: ... behalf of the Toronto Stock Exchange, confirms that as ... no corporate developments that would cause the recent movements ... --> About Aeterna Zentaris Inc. ... --> Aeterna Zentaris is a specialty biopharmaceutical company ...
Breaking Biology Technology:
(Date:10/29/2015)... Oct. 29, 2015  The J. Craig Venter Institute ... "DNA Synthesis and Biosecurity: Lessons Learned and Options for ... Health and Human Services guidance for synthetic biology providers ... --> --> Synthetic ... the potential to pose unique biosecurity threats. It now ...
(Date:10/29/2015)... health pioneer, Joseph C. Kvedar , MD, describes ... wellness, and the business opportunities that arise from it ... Healthy Things . Long before health and wellness ... vice president, Connected Health, Partners HealthCare, was creating a ... the hospital or doctor,s office into the day-to-day lives ...
(Date:10/27/2015)... -- In the present market scenario, security is one ... verticals such as banking, healthcare, defense, electronic gadgets, and ... secure & simplified access control and growing rate of ... bank accounts, misuse of users, , and so on. ... and smartphones are expected to provide potential opportunities for ...
Breaking Biology News(10 mins):