We followed shRNA methods described in our previous report.46 (link) For siRNA, a pool of four siRNA duplexes targeting human or mouse LKB1 and a nontargeting siRNA pool were purchased from Dharmacon, Inc. (Lafayette, CO, USA). Cells were grown in a six-well plate to 40% confluence and incubated with a mixture of a 60 nmol siRNA duplex and 5 μl of transfection reagent 1 (Dharmacon, Inc.). After 4 h, FBS was added to a final concentration of 10% (v/v). Additional transfection was performed at 48-h intervals later for some MEFs.
Sirna duplex
Manufactured by Horizon Discovery
Sourced in United States
SiRNA duplex is a laboratory product used for gene silencing experiments. It is a short, double-stranded RNA molecule that targets and degrades specific mRNA sequences, thereby reducing the expression of a target gene.
Automatically generated - may contain errors
Lab products found in correlation
Sirna duplexes, by Horizon Discovery (1 mentions)
Transfection reagent 1, by Horizon Discovery (1 mentions)
Sigenome library, by Horizon Discovery (1 mentions)
Lipofectamine rnaimax, by Thermo Fisher Scientific (1 mentions)
Pcdna3, by Thermo Fisher Scientific (1 mentions)
Lipofectamine, by Thermo Fisher Scientific (1 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
5 protocols using sirna duplex
1
Knockdown of LKB1 using siRNA
2
Keap1 Silencing Using siRNA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A small interfering RNA (siRNA) duplex targeted against mouse Keap1 and a scrambled, nontargeting control duplex (D-001210-03) were purchased from the Dharmacon siGENOME library. Cells were transfected for 48 h with 10 nM siRNA using Lipofectamine RNAiMAX (Life Technologies), according to the manufacturer׳s instructions.
3
Plasmids and siRNA for ALS Studies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The FLAG-tagged plasmids coding for FLAG-polyGA, FLAG-polyGP, FLAG-polyGR, FLAG-polyPR, and FLAG-polyPA were kindly provided by Prof. Daisuke Ito (Keio University School of Medicine). All plasmids code 100 repeats for each DPRs (Yamakawa et al. 2015 (link)). pCI-HSPB8 plasmid is routinely used in our laboratory and it has been previously described (Crippa et al. 2010b (link); Rusmini et al. 2013 (link)). pcDNA3 (Life Technologies, V790-20) plasmid was used to normalize for transfected plasmid DNA amount. pEGFPN1 (Clontech Lab, U55762) plasmid was used to evaluate transfection efficiency by fluorescent microscopy.
To silence endogenous HspB8 expression, we used a custom small interfering RNA (siRNA) duplex (HspB8 target sequence: CGG AAG AGC UGA UGG UAA AUU; non-target target sequence: UAG CGA CUA AAC ACA UCA AUU) (Dharmacon, Thermo Scientific Life Sciences).
To silence endogenous HspB8 expression, we used a custom small interfering RNA (siRNA) duplex (HspB8 target sequence: CGG AAG AGC UGA UGG UAA AUU; non-target target sequence: UAG CGA CUA AAC ACA UCA AUU) (Dharmacon, Thermo Scientific Life Sciences).
4
Mdm2 Knockdown via siRNA Transfection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For Mdm2 knockdown, the siRNA duplex was designed and custom synthesised from Dharmacon, U.S.A. Mdm2-specific siRNA (5′-CGUACGCGGAAUACUUCGATT-3′) and control siRNA corresponding to luciferase (5′-CCACCUCACAGAUUCCAGCTT-3′) was obtained. HeLa and HEK-293T cells were transfected after every 24 h using Lipofectamine (Invitrogen) with 100 nM of siRNA and evaluated for Mdm2 protein levels. After 48 h of siRNA transfection, cells were transfected with the plasmid of interest using Lipofectamine (Invitrogen).
5
Targeted siRNA Silencing of AMPK
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
siRNA duplex targeting human AMPKα (5′-CCAUACCCUUGAUGAAUUA-3) was purchased from Dharmacon (Lafayette, CO). Transfection of cells with the siRNA duplexes at 50 nM final concentrations was performed using LipofectAMINE 2000 (Invitrogen). AccuTarget™ siRNA (Invitrogen, Carlsbad, CA) was used as a negative control.
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!