Molecular cloning and characterization of StCDPK3

GRANDELLIS C, GIAMMARIA V, ULLOA RM

Mar del Plata

Congreso; XLIII Jornada Anual SAIB; 2007

SAIB

Ca is a crucial signalling molecule in plants involved in thetransduction of environmental, hormonal and developmentalstimuli. Ca -dependent protein kinases (CDPKs) play distinct rolesin mediating Ca signalling. CDPKs are monomeric proteins, withan N-terminal variable domain, a kinase domain, an autoinhibitorydomain and a calmodulin-like domain (CaM-LD).Different CDPK isoforms are expressed during stolon to tubertransition in potato plants: StCDPK3 in early stolons and StCDPK1in induced stolons. was isolated as a partial clonecontaining the autoinhibitory domain, the CaM-LD and part of the3´UTR region; it is 98% homologous to and ,two CDPK from tomato and tobacco. was used to probe agenomic potato library prepared in Dr Glenn Bryan´s lab at SCRI.ABAC clone, B15, containing the gene, was isolated. The5´end of the gene was amplified using primers directed against the5´UTR region of and , subcloned into pGEM-Tand sequenced. StCDPK3 presents an N-terminal myristoylationand palmitoylation consensus. Expression of StCDPK3 wasanalysed in different tissues, the complete coding sequence wascloned and the recombinant StCDPK3-6xHis protein will beexpressed in bacteria. The structure of the gene is being analysed.We expect to obtain promoter sequences to understand thetranscriptional regulation of .