Rt pcr kit

After treatment, cells were lysed in the Trizol reagent (Invitrogen), and RNA was isolated according to the manufacturer’s instructions. The cDNAs for mRNA expression analysis were synthesized from total RNA using RT-PCR kits from Promega (Madison, WI). Real-time qPCR primers are listed in Supplementary Table 3. Some primers used for testing genes related to prostate development were based on a previous report^{51 (link)}.

Total RNA was isolated from cells with TRIzol and cDNA was synthesized using the RT-PCR kit from Promega. Three-step real-time polymerase chain reaction (PCR) was performed using the SYBR Green Mix (BioRad) and an iCycler Iqtm system (BioRad). The primer sequences used for PCR are described in Supplementary Table S1.

Total RNA was extracted from TM4, D1, AR^flox/Y and AR^-/Y;Prrx1::Cre PDCs cells using the TRIzol reagent (Invitrogen) according to the manufacturer’s instructions. cDNA was reversed transcribed from 2 μg mRNA using a Promega RT-PCR kit. For semi-quantitative analyses, PCR was performed on an Applied Biosystems GeneAmp PCR system Veriti 96-well Thermal cycler (27–35 cycles), and band intensities were compared during the linear portion of the amplification cycle. Quantitative PCR was performed on an Applied Biosystems 7500 Real-time PCR detection system using cDNA-specific gene primers and SYBR green master mix (Applied Biosystems). Gene expression levels were compared using the comparative cycle threshold method, with β-actin as input control.
For osteogenesis, microarray analyses were performed on an Applied Biosystems 7500 Real-time PCR detection system using a mouse osteogenesis PCR array kit (Qiagen). The mouse osteogenesis RT² Profiler PCR Array profiles the expression of 84 genes related to osteogenic differentiation. This array contains genes that function in the development of the skeletal system as well as in bone mineral metabolism, and includes growth factors and genes that mediate osteogenesis and related cell growth, proliferation, and differentiation processes. Also represented are ECM molecules and cell adhesion molecules involved in bone development.

To quantify the expression levels of marker genes (PAL4 and NAC4) after PAMP treatments, leaf discs of different transgenic rice plants were immersed in water containing either 100 nm flg22 or 8 nm chitin for 3 h, as described previously (Park et al., 2012). Total RNA was extracted with Trizol reagent (Invitrogen, Cat. No. 15596, Carlsbad, CA, USA) and subjected to first‐strand cDNA synthesis using a Promega RT‐PCR kit (Madison, WI, USA). qRT‐PCR was performed using an iQ5 real‐time PCR detection system (Bio‐Rad). The transcriptional levels were calculated by the relative 2^−ΔCT method with the rice ubiquitin gene as an endogenous reference for normalization.