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:10/11/2017)... ... October 11, 2017 , ... ... Andi Purple announced Dr. Suneel I. Sheikh, the co-founder, CEO and chief research ... Inc. has been selected for membership in ARCS Alumni Hall of Fame ...
(Date:10/11/2017)... (PRWEB) , ... October 11, 2017 , ... Personal eye wash is a basic first ... eye at a time. So which eye do you rinse first if a dangerous substance ... Plum Duo Eye Wash with its unique dual eye piece. , “Whether its ...
(Date:10/11/2017)... ... October 11, 2017 , ... ... will be hosting a Webinar titled, “Pathology is going digital. Is your lab ... digital pathology adoption best practices and how Proscia improves lab economics and realizes ...
(Date:10/11/2017)... LAGUNA HILLS, Calif. , Oct. 11, 2017 /PRNewswire/ ... London (ICR) and University of ... tool to risk-stratify patients with multiple myeloma (MM), in a ... . The University of Leeds is ... Myeloma UK, and ICR will perform the testing services to ...
Breaking Biology Technology:
(Date:4/11/2017)... NXT-ID, Inc. (NASDAQ:   NXTD ) ... appointment of independent Directors Mr. Robin D. Richards ... Directors, furthering the company,s corporate governance and expertise. ... Gino Pereira , Chief Executive ... their guidance and benefiting from their considerable expertise as we ...
(Date:4/5/2017)... 5, 2017 Today HYPR Corp. , ... server component of the HYPR platform is officially ... end-to-end security architecture that empowers biometric authentication across Fortune ... already secured over 15 million users across the financial ... connected home product suites and physical access represent a ...
(Date:4/3/2017)... 2017  Data captured by IsoCode, IsoPlexis ... a statistically significant association between the potency ... and objective response of cancer patients post-treatment. ... whether cancer patients will respond to CAR-T ... as to improve both pre-infusion potency testing and ...
Breaking Biology News(10 mins):