Navigation Links
COS-7

Multiporator Transfection Protocol Protocol No. 4308 915.022 11/1999 Cell line COS-7, monkey, kidney Transfection with Plasmid RSV/lacZ (in bidistilled H2O) Electroporation buffer Eppendorf Hypoosmolar Electroporation Buffer (PH) Culture medium DMEM / 10% FCS Cuvette Eppendorf, 2 mm gap width, 400 l Temperature 4 C Reference Dr. Helena Edlund Department of Microbiology University of Umea Umea Sweden
Phone +46 90 7856701 Fax +46 90 772630 e-mail: Helena.Edlund@micro.umu.se
  1. Harvest the cells in the exponential growth phase and centrifuge them (for 5-10 minutes, 200 x g, at 4 C).
  2. Resuspend the cells in DMEM / 0.5% FCS, determine the number of cells and centrifuge them (for 5-10 minutes, 200 x g, at 4 C). Remove supernatant.

    Note: The overall incubation time in the Eppendorf Electroporation Buffer must not exceed 30 minutes to guarantee a successful electroporation!

  3. Resuspend the cells in Hypoosmolar Electroporation Buffer. When doing so, set the cell concentration to 1 x 106 cells/ml.
  4. Add and mix plasmid DNA (5 g/ml final concentration, in bidistilled H2O).
  5. Transfer 400 l cell suspension into electroporation cuvettes (2 mm gap width). The cell suspension must be free of air bubbles.
  6. Electroporation:

    Mode Eukaryotes Voltage (V) 600 V Time constant (T) 40 s No. of pulses (n) 1
  7. After the pulse, allow the cell suspension to stand in the cuvette for 5 minutes on ice.
  8. Carefully transfer the cell suspension from the cuvette to 3-5 ml DMEM / 10% FCS, and cultivate them in a 55 mm culture dish.
Detection methods for transfection:
The expression of the plasmid RSV/lacZ can be detected after 48 hours with X-gal-staining. Result: Survival rate: n.d. Transfection rate: 30-40% based on the initial number of cells used for the experiment Results were measured 48 hours after transfection.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. A Comparison Study of Lipid Tranfection Reagents in A549, NIH 3T3 and COS-7 Cell Lines
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:6/22/2017)... ... 21, 2017 , ... Building on the success of the inaugural RAADfest last ... very latest developments in radical life extension. RAADfest combines cutting edge science presented for ... of personal development, making it the largest most comprehensive and inclusive super longevity event ...
(Date:6/22/2017)... , ... June 22, 2017 , ... ... results to clients throughout the biopharma and life sciences industries, continue to be ... is seeing. Tunnell’s Kip Wolf will be speaking on “The State of Information ...
(Date:6/20/2017)... ... June 20, 2017 , ... CTNext , Connecticut’s ... has formed a Higher Education Entrepreneurship Advisory Committee to implement the recommendations of ... other high-ranking representatives from 35 higher education institutions across the state over the ...
(Date:6/20/2017)... ... June 20, 2017 , ... National executive search firm, ... extensive assay development and biomarker expertise, as VP of Scientific Affairs at Cambridge ... in bio-analytical assay development and sample testing services. The organization acts as a ...
Breaking Biology Technology:
(Date:4/13/2017)... UBM,s Advanced Design and Manufacturing event in ... and evolving technology through its 3D Printing and Smart ... the expo portion of the event and feature a ... on trending topics within 3D printing and smart manufacturing. ... will take place June 13-15, 2017 at the Jacob K. ...
(Date:4/11/2017)... NXT-ID, Inc. (NASDAQ:   NXTD ) ("NXT-ID" or ... independent Directors Mr. Robin D. Richards and Mr. ... the company,s corporate governance and expertise. ... Gino Pereira , Chief Executive Officer said," ... and benefiting from their considerable expertise as we move forward ...
(Date:4/4/2017)... YORK , April 4, 2017   EyeLock ... today announced that the United States Patent and Trademark ... patent broadly covers the linking of an iris image ... same transaction) and represents the company,s 45 th ... latest patent is very timely given the multi-modal biometric ...
Breaking Biology News(10 mins):