Navigation Links
Saccharomyces cerevisiae

Multiporator / Electroporator 2510 Transformation Protocol Protocol No. 4308 915.531 12/2001 Microorganism Saccharomyces cerevisiae Cell type Yeast Molecules injected Plasmid DNA Growth medium YEPD (1% yeast extract, 2% bactopeptone, 2% dextrose) Washing solution Ice-cold, sterile water; ice-cold 1 M sorbitol Electroporation solution Ice-cold 1 M sorbitol Outgrowth medium 1 M sorbitol Cuvette 2 mm gap width Reference Dr. Robert Sclafani University of Colorado Health Science Center Denver, CO USA Making electrocompetent cells:

1. Inoculate 500 ml YEPD with an aliquot of an overnight culture or a colony from a plate and grow to an O.D.600 of 1.3-1.5 (about 1 x 108 cells/ml). 2. Harvest by centrifugation at 4,000 x g for 5 min. at 4 C. 3. Wash in 500 ml ice-cold sterile water, centrifuge (4,000 x g, 5 min., at 4 C), repeat washing step with 250 ml water. 4. Resuspend in 20 ml ice-cold sorbitol and c entrifuge as above. Resuspend in 0.5 ml of 1 M sorbitol to a final volume of 1.0 to 1.5 ml, keep on ice.

Electroporation of cells:

  1. Add <5 l (0.1 g) DNA to 65 l of electrocompetent cells. Homogenize by gently mixing with pipette several times. Incubate 5 min on ice. 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) 1,500 V Time constant (T) 5 ms
  4. Immediately add 1 ml of cold sorbitol. Plate various aliquots onto selective plates containing 1 M sorbitol.
Expected results: Transformation efficiency up to 104 to 105 transformants/g of DNA. Note: It is possible to scale this protocol down by starting with a culture volume of 50 ml.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Saccharomyces cerevisiae
2. Saccharomyces cerevisiae
3. Saccharomyces cerevisiae
4. Electrophoretic Karyotypes of Wine Strains of Saccharomyces cerevisiae
5. Cytosolic expression of Green Fluorescent Protein (GFP) and its derivatives in the yeast Saccharomyces cerevisiae: Detection in vivo using the Varian Cary Eclipse
6. Monitoring fluorescence resonance energy transfer (FRET) between GFP fusions in lysates of the yeast Saccharomyces cerevisiae using the Varian Cary Eclipse
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:2/11/2016)... , February 11, 2016 ... Corporation ("PositiveID" or "Company") (OTCQB: PSID), a life ... today that its Thermomedics subsidiary, which markets the ... its growth plan in January 2016, including entering ... increasing sequential monthly sales growth, and establishing several ...
(Date:2/10/2016)... - BioAmber Inc. (NYSE: BIOA ), a leader ... & Co. Ltd., its partner in the ... an additional CDN$25 million in the joint venture for ... to 40%.  Mitsui will also play a stronger role ... Sarnia , providing dedicated resources alongside BioAmber,s ...
(Date:2/10/2016)... York, New York (PRWEB) , ... ... ... Pharmaceuticals Inc. (NASDAQ: REGN) today announced that it has joined the Human ... and immunotherapies for infectious diseases and cancer. , The Human Vaccines ...
(Date:2/10/2016)... ... February 10, 2016 , ... HOLLOWAY ... of Pharmaceutical Engineering (ISPE) Rocky Mountain Chapter 21st Annual Vendor Exhibition on Thursday, ... more than 100 tables for its annual event, which will run from 3:00 ...
Breaking Biology Technology:
(Date:2/10/2016)... PUNE, India , February 10, 2016 ... --> According to 2016 iris ... fingerprint identification iris recognition is more widely ... are available with both fingerprint and iris ... allows the user to avoid purchasing two ...
(Date:2/4/2016)... , Feb. 4, 2016 The field ... one of the most popular hubs of the ... and other huge studies of human microbiota, have ... few years, the microbiome space has literally exploded ... research. This report focuses on biomedical aspects ...
(Date:2/2/2016)... 2016  BioMEMS devices deployed in hospitals ... medical screening and diagnostic applications, such as ... that facilitate and assure continuous monitoring without ... being bolstered through new opportunities offered by ... coupled with wireless connectivity and low power ...
Breaking Biology News(10 mins):