Molecular cloning and characterization of a MAPK gene from Polyporus umbellatus

Abstract:

[Objective] To clone the mitogen-activated protein kinase (MAPK) gene from Polyporus umbellatus and carry out the bioinformatics and expression mode analysis. [Methods] RACE technology was carried out to clone the full length cDNA of MAPK gene. The characteristics of physiochemical properties and conserved domains of the predicted MAPK protein were determined using bioinformatics tools. The analyses of multiple alignment and phylogenetic tree were performed using BioEditor and MEGA 5.0 software. Real time quantitative PCR was used for gene expression analysis. [Results] The full length cDNA of MAPK was 1 293 bp in length and encoded a 386-aa protein with a molecular weight of 43.872 kD and an isoelectric point (pI) of 6.68. The PuMAPK clustered with Basidiomycete group according to the phylogenetic analysis. Real time quantitative PCR (qPCR) analysis revealed that transcripts were the most abundant in the beginning of sclerotial formation (20–30 days) with 7.86 fold over that in the mycelium, but the transcripts decreased sharply with the sclerotial development. [Conclusion] Molecular characterization of PuMAPK will be useful for the further functional determination of the gene involving in the development of P. umbellatus sclerotium.

Key words:Polyporus umbellatus, MAPK gene, 5′-RACE, Real-time quantitative PCR, Sclerotium development