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:9/21/2017)... ... September 21, 2017 , ... ... sector professionals, has announced the addition of 5 new courses to its prospectus. ... Compliance with Regulation 21 CFR Part 11 on Electronic Records and Electronic Signatures ...
(Date:9/21/2017)... ... September 21, 2017 , ... Lajollacooks4u welcomed the San Diego chapter of ... worldwide society of professional women with high achievement in the fields of food, fine ... , Twelve members began with an olive oil tasting to whet their palettes ...
(Date:9/20/2017)... ... September 20, 2017 , ... ... with Koch Agronomic Services (Koch) to feature new innovations aimed at helping farmers ... first quarter 2018. American Farmer airs Tuesdays at 8:30aET on RFD-TV. Check your ...
(Date:9/20/2017)... ... September 20, 2017 , ... Diversity focused business accelerator, ... the Growth pitch competition to uncover the top technology-driven, women-led startups in Boston, MA, ... part of each city’s entrepreneurial events going on that week – in Boston, it ...
Breaking Biology Technology:
(Date:4/5/2017)... 2017  The Allen Institute for Cell Science today ... one-of-a-kind portal and dynamic digital window into the human ... first application of deep learning to create predictive models ... and a growing suite of powerful tools. The Allen ... future publicly available resources created and shared by the ...
(Date:4/4/2017)... April 4, 2017   EyeLock LLC , a ... the United States Patent and Trademark Office (USPTO) has ... the linking of an iris image with a face ... represents the company,s 45 th issued patent. ... very timely given the multi-modal biometric capabilities that have ...
(Date:3/30/2017)... The research team of The Hong Kong Polytechnic University ... adopting ground breaking 3D fingerprint minutiae recovery and matching technology, pushing ... for use in identification, crime investigation, immigration control, security of access ... ... A research team led by Dr Ajay ...
Breaking Biology News(10 mins):