pCMV-PKA载体介绍
pCMV-PKA is designed for the constitutive expression of Protein Kinase A (PKA) (mouse) in mammalian cells. It can also be used to study the effects of a given stimulus on the PKA signaling pathway. In conjunction with Clontech’s In Vivo Kinase Assay Kits, pCMV-PKA is an ideal positive control for studying a gene or molecule and its affect on kinase activation in the PKA pathway. cAMP, a ubiquitous second messenger, regulates expression of cellular functions including metabolism, cell proliferation, and neuronal signaling via activation of the PKA-dependent phosphorylation of the CREB transcription factor (1–3). pCMV-PKA contains the gene encoding the catalytic subunit of PKA, which is driven by the human cytomegalovirus (CMV) promoter.
SV40 polyadenylation signals downstream of the PKA gene direct proper processing of the 3’ end of the mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T-antigen. A neomycin resistance cassette (Neor)—consisting of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the Herpes simplex virus thymidine kinase (HSV TK) gene—allows stably transfected eukaryotic cells to be selected using G418 (4). A bacterial promoter upstream of this cassette expresses kanamycin resistance in E. coli. The pCMV-PKA backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production. The recombinant vector can be transfected into mammalian cells using any standard method.
