Navigation Links
Agrobacterium tumefaciens

Multiporator / Electroporator 2510 Transformation Protocol Protocol No. 4308 915.502 12/2001 Microorganism Agrobacterium tumefaciens Cell type Bacteria, gram negative Molecules injected Plasmid DNA (in water) Growth medium TB medium Washing solution Sterile, cold water Electroporation solution 10% glycerol Outgrowth medium LB medium Cuvette 1 mm gap width Reference Mersereau M., et al 1990 Gene 90 149-151

Making electrocompetent cells:

  1. Grow cells at 30 C to a density of O.D.600: 1.5-2.0.

  2. Harvest by centrifugation and wash five to seven times in sterile, cold water.

  3. Resuspend the cells in 10% glycerol to a final concentration of 4-6 x 1011 cell/ml and store frozen at 70 C.

  4. For electroporation frozen cells were thawed in ice and diluted with sterile water to a density of 1 x 1010 cells/ml prior to use.

Electroporation of cells:

  1. Add 2 ng plasmid DNA to 40 l of electrocompetent cells. Homogenize by gently mixing with pipette several times.

  2. Transfer mixture into a prechilled cuvette.
  3. Wipe moisture from the cuvette and insert the cuvette into the device.

  4. Electroporation:


  5. Mode Prokaryotes O Voltage (V) 1440 V Time constant (T) 5 ms
  6. Immediately add 400 l LB medium and transfer into a sterile tube. Incubate at room temperature for 1 hour.
  7. Plate cells on minimal selective AB plates; incubate 12-72 hours.

Expected results:

Transformation efficiency up to 1-3 x 108 transformants/g of DNA.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Agrobacterium tumefaciens
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:6/27/2016)... Hill, N.C. (PRWEB) , ... June 27, 2016 ... ... U.S. commercial operations for Amgen, will join the faculty of the University ... serve as adjunct professor of strategy and entrepreneurship at UNC Kenan-Flagler, with a ...
(Date:6/24/2016)... Epic Sciences unveiled a liquid biopsy ... PARP inhibitors by targeting homologous recombination deficiency (HRD) ... test has already been incorporated into numerous clinical ... Over 230 clinical trials are investigating ... PARP, ATM, ATR, DNA-PK and WEE-1. Drugs targeting ...
(Date:6/23/2016)... ... June 23, 2016 , ... UAS LifeSciences, one of ... their brand, UP4™ Probiotics, into Target stores nationwide. The company, which has been ... Target to its list of well-respected retailers. This list includes such fine stores ...
(Date:6/23/2016)... Houston Methodist Willowbrook Hospital has signed ... to serve as their official health care provider. ... will provide sponsorship support, athletic training services, and ... volunteers, athletes and families. "We are ... and to bring Houston Methodist quality services and ...
Breaking Biology Technology:
(Date:6/20/2016)... 20, 2016 Securus Technologies, a leading ... for public safety, investigation, corrections and monitoring announced ... it has secured the final acceptance by all ... Managed Access Systems (MAS) installed. Furthermore, Securus will ... be installed by October, 2016. MAS distinguishes between ...
(Date:6/9/2016)... , June 9, 2016 ... Police deploy Teleste,s video security solution to ensure the safety ... France during the major tournament Teleste, ... communications systems and services, announced today that its video security ... to back up public safety across the country. ...
(Date:6/2/2016)... --  The Weather Company , an IBM Business (NYSE: ... in which consumers will be able to interact with IBM ... voice or text and receive relevant information about the product ... have long sought an advertising solution that can create a ... and valuable; and can scale across millions of interactions and ...
Breaking Biology News(10 mins):