Dlx-2 siRNA (h)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Use si-DLX2: 50 μM/2.5×10^5 cells. Incubate the plate at 37C in a CO2 incubator for 4 hours. Change the medium and continue incubation for 24 hours.	In case of high toxicity, reduce the amount of DNA and/or HilyMax

Protocol tips
Use si-DLX2: 50 μM/2.5×10^5 cells. Incubate the plate at 37C in a CO2 incubator for 4 hours. Change the medium and continue incubation for 24 hours.
Downstream tips
In case of high toxicity, reduce the amount of DNA and/or HilyMax

Publication protocol

Construction of the DLX2 overexpression vector and transfection
An insert of human DLX2 was amplified from pCMV-sports6/DLX2 (Korea Human Gene Bank, Seoul, Korea) by PCR using the following primers: EcoRI (forward): 5'-CGGAATTC ATGACTGGAGTCTTTGAC-3' and XhoI (reverse): 5'-CTCTGAGT TTAGAAAATCGTCCCCG-3'. DLX2 inserts was cloned in the vector pcDNA3-myc which allows expression of c-myc-tagged protein in mammalian cells. pcDNA3-myc/DLX2 or pcDNA3-myc was then delivered to cells (4 μg/2.5 × 105 cells) using HilyMax (Dojindo Molecular Technologies, Rockville, ML, USA) transfection reagent according to the manufacturer’s instruction. After incubation for 24 h, the cells were detached and irradiated.

Small-interfering RNA (siRNA) transfection
The si-RNAs targeting DLX2 and Smad2/3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). For transfection, cells were plated and grown to 70–90% confluence. Target si-RNA or negative control si-Ct was then delivered to cells (si-DLX2: 50 μM/2.5×105 cells; si-Smad2/3: 100 μM/2.5×105 cells) using HilyMax transfection reagent according to the manufacturer’s instruction. After incubation for 24 h, the cells were detached and irradiated.

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Manufacturer protocol

Download the product protocol from Santa Cruz Biotechnology for Dlx-2 siRNA (h) below.

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