UALH were generated by transfection of osteocarcinoma cancer U2OS cells with the (ARE-RAD9)6-collagenase-Luciferase-SV40-hygromycin plasmid.
UALH hAR cells were generated by co-transfection of UALH cells with the plasmids pSG5-hAR-puromycin and pSG5-hAR-Renilla-luciferase-(ARE-RAD9)6-collagenase-Luciferase-SV40-neomycin.
The plasmid (ARE RAD9)6-Coll-Luc-SVNeo contains a luciferase gene driven by the mouse mammary tumor virus promoter and a neomycin phosphotransferase gene under the control of SV40 promoter.
The pSG5-puro plasmid pSG5-human AR-puromycin enables to express the human androgen receptor (hAR). Puromycin N-acetyl transferase selection marker expression confers resistance to puromycin.
The plasmid pSG5-hAR-Renilla-Neomycin contains the gene coding for hAR under the control of the SV40 promoter, the Renilla luciferase reporter gene under the control of the constitutive human cytomegalovirus (CMV), the firefly luciferase gene under the control of six androgen response elements (ARE) from the RAD9 gene promoter and placed upstream of the collagenase promoter, and the resistance gene to neomycin under the control of the SV40 promoter. This third plasmid has been included to over-express both AR and luciferase expression.