ON-TARGETplus SMARTpool siRNAs targeting Arid5a, HuR, TRAF2, and MCPIP1 were from Dharmacon (Lafayette, CO). For RNA silencing, ST2 cells and MEFs were seeded overnight in antibiotic-free α-MEM, and N/TERT2G cells were seeded in KC serum-free media (Gibco) with supplements. Transfection was performed 18 hours later with 50 nM siRNAs with DharmaFECT Reagent 1. Culture media were replaced after 24 hours, and IL-17 was administered 24 hours later. Flag/Myc-Arid5a was from OriGene (Rockville, MD). All other constructs were described (6, 22, 27, 63). Transfections of ST2 cells and MEFS were performed with FuGENE HD (Promega, Madison, WI) or CaPO4 (HEK293T cells). RNA was prepared with RNeasy kits (Qiagen, Valencia, CA). Complementary DNA was synthesized by SuperScript III First Strand Kits (Thermo Fisher Scientific, Waltham MA). Quantitative real-time PCR (qPCR) was performed with the SYBR Green FastMix (Quanta BioSciences, Beverly, MA) and analyzed on an ABI 7300 real-time instrument. Primers were from QuantiTect Primer Assays (Qiagen).

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.