Molecular Cloning Of Polyketide Synthase Genes From Monascus Aurantiacus

Fungal polyketides are a large family of secondary metabolites exhibit a vastdiversity of structure and function, including lovastatin, pigments and mycotoxins.The biosynthesis of fungal polyketides is governed by key enzymes. In this paper, wecloned PKS genes from Monascus aurantiacus AS3.4384, to make the foundation ofthe characteration of function and the cloning of full-length PKS genes. The mainresearch results are as follows:1 Two pairs of degenerate primers (pksF1/R1 and pksF1/R2) were designedbased on sequences from known fungal PKS genes. Four PKS fragments wereamplified from Monascus aurantiacus AS3.4384 using pksF1/R1 and pksF1/R2, andnamed pks1, pks2, pks3 and pks10, respectively. The Blast searches (NCBI) revealed,pks2 was a part of pks1 (7144bp, accession nos. AJ414729 in GenBank) in Monascuspurpureus, reported only in a database.2 Three specific primers (PF1/PR1, PF3/PR3 and PF10/PR10) were designedbased on sequences of pks1, pks3 and pks10, respectively. Three positive fosmidclones were screened from a fosmid library of Monascus aurantiacus AS3.4384 usingeach of the three specific primer pairs. After distruption of positive fosmids byultrasonication, the sonicated DNA were subcloned into pUC18 to construct three2～4Kb libraries. Three PKS genes were obtained from the corresponding subclonelibrary, and designated MApks1, MApks3 and MApks10, respectively.3 By bioinformatics analysis, the deduced amino acid sequence of MApks1 waspossiblely the citrinin biosynthetic PKS and classified as non-reducingⅢPKS,whereas the translated sequences from MApks3 and MApks10 had homologous withPKS involved in T-toxin and lovastatin biosynthesis and classified as reducing PKS.