Lipofectamine 2000 protocol pdf

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Lipofectamine 2000 protocol pdf

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