For biochemical experiments, HEK293 cells were individually transiently transfected with the following plasmids: 240 ng of HA-TRPM4 WT or HA-TRPM4 N992Q, 2 μg of either TRPM5 WT or TRPM5 N932Q in a P100 dish (BD Falcon, Durham, North Carolina, USA), mixed with 30 μ l of JetPEI (Polyplus transfection, Illkirch, France) and 250 μ l of 150 mM NaCl. The cells were incubated for 48 h at 37°C with 5% CO2. For electrophysiological studies, T25 cm2 flasks of HEK293 cells were transiently co-transfected using X-tremeGENE 9 DNA transfection mix reagent (Roche Diagnostics, IN, USA) with 0.3 μg of WT, mutant TRPM4, or TRPM5 channels. These transfections included 0.2 μg of cDNA encoding CD8 antigen as a reporter gene. Anti-CD8 beads(Dynal, Oslo, Norway) were used to identify transfected cells, and only CD8-displaying cells were analyzed.Cells were used 48 h after transfection.
X tremegene 9 dna transfection mix reagent
Manufactured by Roche
Sourced in United States
X-tremeGENE 9 DNA transfection mix reagent is a cationic lipid-based transfection reagent designed for efficient delivery of DNA into a variety of cell lines. It facilitates the uptake of DNA into the cells, enabling genetic modification and gene expression studies.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using x tremegene 9 dna transfection mix reagent
1
TRPM4 and TRPM5 Channel Assays
2
Transient Transfection of TRPM4 Variants in HEK293
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human embryonic kidney (HEK293) cells were cultured with DMEM supplemented with 4 mmol/L glutamine, 10% FBS, and 20 μg/mL gentamycin. They were transiently transfected with 240 ng of HA‐TRPM4 wild type (WT), variants (D198G, A432T, G582S, A432T/G582S, T677I and V921I), or empty vector (pcDNA4TO) in a P100 dish (BD Falcon), mixed with 30 μL of JetPEI (Polyplus‐transfection) and 250 μL of 150 mmol/L NaCl. The cells were incubated for 48 hours at 37°C with 5% CO2. The amount of cDNA used was proportionately increased to the amount used for electrophysiological studies. For electrophysiological studies, T25 25‐cm2 flasks of HEK293 cells were transiently cotransfected using X‐tremeGENE 9 DNA transfection mix reagent (Roche Diagnostics) with 80 ng WT or variant TRPM4 channels. All transfections included 200 ng cDNA encoding CD8 antigen as a reporter gene. Anti‐CD8 beads (Dynal) were used to identify transfected cells, and only CD8‐displaying cells were analyzed. Cells were used 48 hours after transfection.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!