Lipofectamine 2000 protocol pdf
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nucleic acid- lipofectamine® complexes can be added directly to cells in culture medium, in the presence or absence of serum/ antibiotic. sirna/ chimera transfection. the reagent allows for transfection to be performed both in presence or absence of serum. add the oligomer- lipofectamine complexes to each well containing cells and medium. successful transfection every time unmatched versatility, high efficiency with transfection success across multiple cell lines, lipofectamine™ gives you the most versatile route to. 75 μl of lipofectamine ltxtm reagent into the above diluted opti- mem® : dna solution, mix gently and incubate 30 minutes at room temperature to form dna- lipofectamine ltxtm reagent. aspirate off pbs 5. add 50 μl of dna to the diluted transfection reagent from step 2. add 1 μg of plasmid dna into each of the tubes labeled 3: 1 and 6: 1, and 2 μg of dna ( also in a total of lipofectamine 2000 protocol pdf 50 μl) into the tube labeled 3: 2. after the 5- minute incubation, combine the diluted oligomer with the diluted lipofectamine. lipofectamine™ pdf reagent is sufficient for 500 to 1, 000 transfections in 24- well plates, ortransfections using 6- well plates. add 2- 3 ml pbs to wash 4. lipofectamine® transfection reagent is a proprietary formulation for transfecting nucleic acids ( dna, rna, and mrna) into a wide range of eukaryotic cells. incubate 30min at 37° c while mixing following components in hood ( see spreadsheet for amounts, this example is for one 10cm. for each well of cells to be transfected, dilute 0. dna- lipofectamine® complexes must be made in serum- free medium such as opti- mem® reduced serum medium and can be added directly to cells in culture medium, in the presence or absence of serum. product lipofectamine® reagent is a proprietary formulation description for transfecting nucleic acids into a wide range of eukaryotic cells. add 8 ml of 10% fbs/ opti- mem media / plate 6. sirna/ chimera should be stored at 4° c and at - 20° c in case of dried and soluble forms respectively, but should not be refrozen for reuse. lipofectamine transfection manufacturer protocol day 0: lipofectamine 2000 protocol pdf seed cells onto the wellplates / petridishes to be transfected so that day 1 is at 70- 80% confluence day 1: transfection • vortex and centrifuge dna epps • prepare mix a and mix b for each sample and incubate mixtures separately for 5 min o mix a 6 wellsplate well 10 cm dish. the easy protocol, exceptional plasmid dna delivery, and the ability to. 5 μg of dna in 100 μl of opti- mem® i reduced serum media without serum. it is not necessary to remove complexes or. mix gently and incubate for 20 minutes at room temperature ( solution may appear cloudy). aspirate medium off cells 3. with pdf lipofectamine. researchers use lipofectamine™ reagent for sirna- and shrna- based gene knockdown experiments, as well as for gene expression studies. for each well of cells, add 0. tap the tubes or vortex for 1 sec to mix the contents and incubate for 15 – 45 min at room temperature. lipofectamine™ transfection reagent is a versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines. to optimize the amount of lipofectamine™ for transfection in a 24- well plate, start with cells at > 90% confluency and use a fixed amount of dna ( 0. mix gently by rocking the plate back and forth. ( 10 2000 μl) should be made. with cell number and dna concentration held constant, vary the amount of lipofectamine™ to determine the optimal concentration ( usually 2000 1. efficient rnai can be achieved by using commercially available trasnfection reagent such as lipofectamine and lipofectamine rnaimax. make sufficient 10% fbs in opti- memmedia ( w/ oantibiotics) sterile filter 2.