Fugene is a transfection reagent that apparently increases transfection rates for animal cells. Lfugene hd transfection reagent remains fully functional even. You use the optimized reagents from the lightswitch luciferase assay system including the lightswitch luciferase assay reagents and fugene hd for best results. Fugene hd transfection reagent is a multicomponent reagent that forms a complex. Detecting recruitment of target proteins to the proteasome in. The fugene hd to plasmid dna ratio shown in this example is 3. Fugene hd transfection reagent quick protocol fb112pdf. Detecting recruitment of target proteins to the proteasome. To be the most responsive supplier of biological reagents and reagent systems used in research and applied technology applications worldwide. The reagent is stable thro ugh the expiration date printed on the label when stored under these conditions. Fugene6, and fugenehd are trademarks of fugent, llc.
Search our fugene hd protocol database if youre new to the reagent or exploring the use of a new cell line. In animal cells, transfection is the preferred term as. Briefly vortex the xtremegene hp dna transfection reagent vial. The protocol does not require removal of serum or culture medium and does not require washing or changing of medium after introducing the reagentdna complex. Many times, a scientist may need to transfect different constructs or even different molecules e. The protocol does not require removal of serum or culture medium and does not. Protocol 2 producing lentivirus in hek293t cells using a. The use of fugene hd does not require culture medium change or serum removal during. Follow the transfection reagent protocol for seeding cells. Fugene is a registered trademark of fugent, llc in the united states and other countries. Lipofectamine stem transfection reagent thermo fisher. Improving the success of your transfection promega connections.
It is suitable for use in media with or without serum, and for transient or stable transfection, as well as cotransfections of multiple dna plasmids. Always mix fugene 6 reagent prior to use vortex for one second or use inversion. Not every lab has a tried and true transfection protocol that can be used by all lab members. Fugene hd is added, the transfection mix is vortexed for 1 s, and then incubated for 15 min. Aug 30, 2019 not every lab has a tried and true transfection protocol that can be used by all lab members. To a sterile tube or u or vbottom plate, add 9098l of medium prewarmed to room temperature so that the final volume after. Fugene 6 transfection reagent is stabilized for extended storage. Fugene 6 transfection reagent is a nonliposomal formulation designed to transfect plasmid dna into a wide variety of cell lines with high efficiency and low toxicity.
Fugene hd transfection reagent promega corporation. Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. The amount of dna and the fugene hd transfection reagent in protocol described bellow calculated to be sufficient to transfect cells in 20 wells of 96well. In 6cmdish, when the confluence of 293t reachs 50%,i conduct the transfection. Find the right product for your cells by using our selection guide. Description fugene hd transfection reagent is a proprietary, nonliposomal, low toxicity formulation designed for high efficiency delivery of dna into a broad range of cell lines. For specific goclone transfection protocol examples, see our online.
Fugene hd transfection reagent for efficient mammalian cell. The protocol does not require removal of serum or culture. This reagent is suitable for use in media with or without serum, and for transient or stable transfection. Fugene 6 transfection reagent is gentle on the cells. Additionally, low cell numbers can be transfected in 96well plates. Replating transfected hek293 cells into multiwell plates and adding halotag ligand 1. The protocol can be scaled to produce different amounts of virus as needed. Dilute dna with appropriate diluent, such as serumfree medium to a final concentration of 1.
Fugene hd transfection reagent for efficient mammalian. Fugene hd transfection reagent is a proprietary, nonliposomal, low toxicity formulation designed for high efficiency delivery of dna into a broad range of cell lines. Add transfection mixture to wells with attached cells and express proteins overnight 1824 hours at 37c, 5% co 2. Few researchers will use the same cell type and same construct to generate data. Transfection mixes are added dropwise to each well of the 6well plate, and the cells are incubated for 48 h in a humidified atmosphere at 37 c5% co 2.
Jul 15, 2016 the fugene hd transfection reagent protocol database has protocols for transfecting your cell line. Adherent cells can be trypsinized and transfected by the dna. The fugene hd transfection reagent is a nonliposomal formulation designed to transfect dna into a wide variety of cell lines with high efficiency and low toxicity. Fugene hd transfection reagent offers unrivaled transfection performance with minimal impact on cell physiology jacobson et al. Optimize transfection of cultured cells promega corporation. Add transfection mixture to wells with attached cells and express proteins for approximately 20 hours at 37c, 5% co 2. Hek293 cells were grown to 85% confluency, then harvested and plated in a 96well plate at 2. Detecting ubiquitination of target proteins in live cells. Fugene 6 transfection reagent is a proprietary blend of lipids and other components supplied in 80% ethanol, sterilefiltered, and packaged in glass vials. Before attempting this protocol, please make sure that. The use of fugene hd does not require culture medium change or serum removal during transfection experiment. Before the experiment, i place the optimem at room temperature for 10min and vortex the fugene 6 to make it evenly. Transfection of sf9 cells in suspension nih common fund. Use 2g dna and 4l fugene per 10cm dish scale this up to prepare in bulk.
Cultures were observed over 48 hours from start of transfection. Vwr offers transfection reagents for numerous applications, including bioproduction, in vivo work, viral production, dna and sirna transfection, crisprcas 9 and more. Add transfection mixture to wells with attached cells and. Promega corporation is a worldwide leader in applying biochemistry and molecular biology to the development of innovative, highvalue products for the life sciences. Add 6l of fugene hd transfection reagent and incubate at room temperature for 10 minutes. Has an easily optimizable protocol that allows it to transfect a wide range of eukaryotic cells, including animal and insect cells, with minimal cytotoxicity. The lipofectamine ltx and fugene hd lipidbased transfection reagents were.
A protocol for nonliposomal transfection of dna into a variety of cell lines with high efficiency and low toxicity. Detecting the interaction of target proteins with e3. Fugene hd transfection reagenta is a novel, nonliposomal formulation designed to transfect dna into a wide variety of cell lines with high efficiency and low toxicity. Fugene hd transfection reagent is a multicomponent reagent that forms a complex with dna, then transports the complex into animal or insect cells. These protocols guide the transfection process for many cell lines, and will also calculate the volumes needed based on well format and the number of samples. Fugene is apparently a nonliposomal formulation of blended lipids, but its actual contents are not revealed anywhere. Additionally, the fugene hd transfection reagent has been shown to support transfection in chemically defined. This is the preferred protocol for use in highthroughput screening applications. Tips and tricks on how to improve transfection experiments. Promega corporation 2800 woods hollow road madison, wi 537115399 usa toll free. The day before transfection plate adherent cells in the amount that by the time of transfection.
Transfection protocol fugene hd transfection reagent instructions for use of products e2311 and e2312. Cells were visualized 48 hours posttransfection and imaged. L overview fugene hd transfection reagent is a multicomponent reagent that forms a complex with dna, then transports the complex into animal or insect cells. Storage and stability fugene 6 reagent is shipped at room temperature. Select the cell line, type of plate used and number of wells to be transfected and a protocol will be generated based on conditions tested by promega scientists. Fugene 6 reagent complex prior to plating, making it a strong candidate for high throughput applications. Add 3 ul fugene hd directly to the transfection medium, avoid touching the side of the tube when adding fugene. Fugene transfection protocol 1 add 3ul of fugene to 97ul of optimem, being sure to avoid touching the pipette tip to the side of the tube as the fugene will stick to plastic. We use it in lab for cell culture transfections, but i have no idea what it is or how it works.
Optimal transfection conditions for hek293 with the fugene hd reagent were determined using the protocol recommended in figure 2. Fugene hd transfection reagent quick protocol fb112pdf 99 kb english. Fugene hd transfection reagent protocol promega corporation. To a sterile tube or u or vbottom plate, add 90 98l of medium prewarmed to room temperature so that the final volume after. Fugene hd transfection reagent manual active motif. A protocol for nonliposomal transfection of dna into a variety of cell lines with high efficiency and low. Xtremegene hp dna transfection reagent sigmaaldrich.
Improving the success of your transfection promega. All three offer broad spectrum transfection capability and low. It may also refer to other methods and cell types, although other terms are often preferred. Protocol 2 producing lentivirus in hek293t cells using a 2.
Promega offers three premium transfection reagents. These protocols were developed by promega corporation or fugent, l. Description fugene 6 transfection reagenta is a nonliposomal reagent that transfects dna into a wide variety of cell lines with high e. This reagent is suitable for use in media with or without serum. Transitlt1 transfection reagent research reagent products. Gently drip 5l or 3l 96 or 384well formats onto seeded cells. Select the cell line, type of plate used and number of wells to be transfected and a protocol will be. Description the nanobret cmycmax interaction assayah monitors the interaction between human cmyc and max in living cells. While considering a suitable transfection reagent, it is. Fugene hd transfection reagent is a nonliposomal formulation designed. Transfect cells according to the manufacturers protocol. Utilizes a protocol that does not require removal of serum or culture medium and does not require washing or changing of medium after introducing the reagentdna.
Fugene hd transfection reagent is a novel, nonliposomal formulation designed to transfect dna into a wide variety of cell lines with high efficiency and low toxicity. Fugene hd transfection reagent lets you work in your cancer model. Xtremegene transfection reagents from roche have been used for a wide range of applications and a vast array of cell lines, primary cells and stem cells. Promega fugene hd transfection reagent fugene hd is a novel, nonliposomal reagent for transfecting dna into a variety of cell lines with high efficiency and low toxicity. Use the fugene hd protocol database to see if transfection conditions for your cell line are available. Transfection of j774 macrophages using fugene hd from roche based on preliminary experiments conducted by barish ozdamar in fred maxfields lab, fugene hd from roche consistently gives 510%. The fugene hd protocol database is a collection of protocols that guide the transfection process for a variety of cell lines. Perform transfection into hek293t as above using fugene hd roche applied bioscience and harvest these cells 48 h later for use in.
452 1073 1504 701 1179 1222 405 442 1574 402 350 31 1033 188 556 1365 250 1 375 1375 822 1147 671 523 983 468 529 467 306 319 1452 1495 494 1454