Navigation Links
Primary cells, rat heart muscle

Multiporator Transfection Protocol Protocol No. 4308 915.035 12/1999 Cell line Primary cells: Adult heart muscle cells from rat, 6 days in culture Transfection with Plasmid pCMV--galactosidase (in bidistilled H2O) Electroporation buffer Eppendorf Isoosmolar Electroporation Buffer (PI) Culture medium M199 / HEPES (2 mM carnithin, 5 mM taurine, 10 M cytosine-arabinofur.)+ 20% FCS Outgrowth medium HEPES-buffer (140 mM NaCl, 3.6 mM KCl, 1.2 mM MgSO4, 1 mM CaCl2, 11 mM glucose, 100 U/ml penicillin, 100 g/ml streptomycin, 20 mM HEPES, pH 7.4) without phenol red + 10% FCS Cuvette Eppendorf, 2 mm gap width, 400 l Temperature RT (20-25 C) Reference Dr. Gerhild Taimor Justus-Liebig-Universitt Physiologisches Institut Aulweg 129 D-35392 Gieen
Phone +49 641 9947243 Fax +49 641 9947239 mail: gerhild.taimor@physiologie.med.uni-giessen.de
  1. Harvest the cells by collagenase treatment (0.4 mg/ml, 30 min., 37 C) and centrifuge them (for 2 minutes, 300 x g, at room temperature).
  2. Resuspend the cells in M199 / 0.5% FCS, determine the number of cells and centrifuge them (for 2 minutes, 300 x g, at room temperature). 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 Isoosmolar Electroporation Buffer. When doing so, set the cell concentration to 1 x 104 - 105 cells/ml.
  4. Add and mix plasmid DNA (5-20 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) 100 V Time constant (T) 50 s No. of pulses (n) 1
  7. After the pulse, allow the cell suspension to stand in the cuvette for 10 minutes at room temperature.
  8. Carefully transfer the cell suspension from the cuvette to 3-5 ml HEPES medium without phenol red / 20% FCS, and cultivate them in a 35 mm culture dish.

Detection methods for transfection:

The expression of the plasmid pCMV--galactosidase can be detected clearly after 24-48 hours with X-gal-staining under a microscope.


'"/>

Source:


Page: All 1 2

Related biology technology :

1. Human Primary Preadipocytes and Differentiated Adipocytes
2. Primary cells, rat heart endothel
3. Efficient Delivery of siRNAs to Human Primary Cells: Electroporation vs. Chemical Transfection
4. Deliver siRNAs Into Primary Cells
5. High Throughput siRNA Delivery In Vitro: From Cell Lines to Primary Cells
6. Electroporation of Primary Bone Marrow Cells
7. Efficient Primary Structure Elucidation of Disulfide-bridged Peptides
8. Embryonic stem cells, mouse
9. Laser Capture Microdissection of muscle fiber populations and expression analysis by RT-PCR
Post Your Comments:
*Name:
*Comment:
*Email:
TAG: Primary cells rat heart muscle

(Date:1/22/2015)... PALO ALTO, Calif. , Jan. 22. 2015  Varian Medical ... equipment and software, has been honored for its commitment to ... most sustainable companies. Varian is the highest ranked healthcare equipment ... ranking, announced today during the World Economic Forum at ...
(Date:1/22/2015)... 22, 2015  Derma Sciences, Inc. (Nasdaq: ... advanced wound care, announces that AMNIOEXCEL® and AMNIOMATRIX®, ... added to the Premier, Inc. Regenerative Skin Grafting ... the AMNIOEXCEL® and AMNIOMATRIX® product lines, which continue ...
(Date:12/24/2014)...  United Therapeutics Corporation (NASDAQ: UTHR ) announced ... has submitted a pre-market approval application to the U.S. ... Medtronic,s SynchroMed ® II implantable drug infusion system ... Therapeutics, Remodulin ® (treprostinil) Injection delivered intravenously to ...
(Date:12/24/2014)... Dec. 24, 2014  Vermillion, Inc. (Nasdaq:   VRML), ... announced today that the Company closed its previously ... Management, Jack W. Schuler , Birchview Fund ... $10.5 million, before offering expenses.  The proceeds will ...
Breaking Biology Technology:Varian Honored Among World's 100 Most Sustainable Corporations 2Derma Sciences Expands Access of its AMNIOTIC TISSUE Product Line with New Premier, Inc. Agreement 2Derma Sciences Expands Access of its AMNIOTIC TISSUE Product Line with New Premier, Inc. Agreement 3Derma Sciences Expands Access of its AMNIOTIC TISSUE Product Line with New Premier, Inc. Agreement 4Derma Sciences Expands Access of its AMNIOTIC TISSUE Product Line with New Premier, Inc. Agreement 5United Therapeutics Announces Submission Of Pre-Market Approval Application For Implantable Drug Infusion System To Deliver Remodulin 2United Therapeutics Announces Submission Of Pre-Market Approval Application For Implantable Drug Infusion System To Deliver Remodulin 3United Therapeutics Announces Submission Of Pre-Market Approval Application For Implantable Drug Infusion System To Deliver Remodulin 4Vermillion Announces Closing of Equity Financing 2Vermillion Announces Closing of Equity Financing 3
... Calif., March 12 ,Exelixis, Inc. (Nasdaq: EXEL ) ... executive officer of Exelixis, will present at the,Cowen and ... / 1:00 p.m.,PT on Monday, March 17, 2008. Dr. ... corporate strategy., The presentation will be webcast and ...
... DIEGO, March 12 Biomatrica, the leader in ... a reagent that enhances PCR and RT-PCR performance ... amplification of genomic,DNA or RNA templates., PCRboost ... that are,traditionally difficult to amplify and present in ...
... 12 Oncolys BioPharma,Inc. (Tokyo, Japan) and Tacere ... they have entered into a strategic alliance and,license ... interference (RNAi)-based Hepatitis C virus (HCV) compound, TT-033,code-named ... the,strategic alliance entered into by Tacere and Oncolys ...
Cached Biology Technology:Biomatrica's PCRboost(TM) Enables 5-Fold Improvement in Amplification of Difficult Samples 2Oncolys BioPharma and Tacere Therapeutics Sign Strategic Alliance and License Agreement to Develop and Commercialize RNAi Hepatitis C Drug 2Oncolys BioPharma and Tacere Therapeutics Sign Strategic Alliance and License Agreement to Develop and Commercialize RNAi Hepatitis C Drug 3
(Date:1/22/2015)... Jan. 15, 2015  BellBrook Labs, a leader in ... the launch of a TR-FRET (time resolved Forster ... UDP Assay, a high throughput screening assay for ... will allow for sensitive detection of hundreds of ...
(Date:1/22/2015)... , Jan. 21, 2015  Analyst Report Issued by Small Cap ... report showed that planet-wide, transactions at merchants on the leading ... total number of credit, debit, and prepaid cards reached 6.54 ... victims of fraudulent card usage in 2012, and credit and ...
(Date:1/22/2015)... CHARLOTTE, N.C. , Jan. 22, 2015 Infinisource has ... introduction of the G2 model. The G2 sets a higher ... company,s iSolved human capital management solution. With plug-and-play installation, touch ... G2 time clock provides a robust time collection solution for ...
Breaking Biology News(10 mins):BellBrook Labs Launches a High Throughput Screening Assay for Glycosyltransferases with TR-FRET Readout 2$4.80 Near Term Price Target with "Buy" given by CapRock Research for NXT-ID, Inc. (NASDAQ: NXTD) issued by Small Cap IR 2Infinisource's NXG series sets new time clock standard 2
... Ridge Associated Universities (ORAU), a consortium of 98 Ph.D.-granting ... sciences in the University of Delaware,s College of Earth, ... Junior Faculty Enhancement Award. The competitive award, which ... from the faculty member,s university, is intended to enrich ...
... the National Oceanography Centre (NOC) joins forces with the ... Porcupine Abyssal Plain sustained observatory (PAP-SO), the longest multidisciplinary ... advance scientific understanding of the ocean and improve climate ... North Atlantic off the Irish continental margin, where the ...
... PITTSBURGH, May 25 A two-year, $12 million contract ... Transition (OTT) will jumpstart human trials of three innovative ... with healthy, functional tissues, announced officials of the University ... today at the Institute,s Second Annual Open Session, Soldiers ...
Cached Biology News:University of Delaware professor wins Powe Award from Oak Ridge Associated Universities 2University of Delaware professor wins Powe Award from Oak Ridge Associated Universities 3Met Office and NOC enhance ocean observatory 2Pitt gets $12 million DoD contract for regenerative medicine treatment trials 2
Request Info...
Prepared with distilled water. Yeast extract solution is autolyzed at 150 g/L....
Streptomycin sulfate an aminoglycoside used for bacterial selection binds to the small ribosomal subunit and interferes with protein biosynthesis.21H39N7O12)2x3H2SO4Formula weight:...
...
Biology Products: