title: Bamboozled again! Inadvertent isolation of fungal rDNA sequences from bamboos (Poaceae: Bambusoideae).
abstract: Polymerase chain reaction (PCR) amplification of the nuclear internal transcribed spacer (ITS) and 5.8S regions of rDNA from woody bamboos (Bambuseae) led to the recovery of fungal instead of bamboo sequences under a variety of PCR conditions and irrespective of whether the plant DNA was extracted from fresh leaves or silica gel-dried material. Phylogenetic analyses based on the 5.8S sequences indicated that the fungi were most likely basidiomycetes and that none was an ascomycete. A diverse assemblage of nonascomycetous fungi was isolated from different bamboos, and various fungi coexisted in the same host plant. There was no evidence that closely related fungi consistently associate with closely related host bamboos. Phylogenetic analysis based on 5.8S sequences showed that some fungi were in lineages near Volvariella, Lentinula, Peniophora, and Rhizoctonia, but the insufficiency of basidiomycete and zygomycete ITS sequences in sequence data bases precluded more precise fungal identifications. Bamboo ITS regions were amplified only when fresh leaves were surface sterilized before DNA extraction, suggesting that the fungal associates are epiphyllous rather than endophytic. This study highlights the possibility of inadvertent PCR amplification of contaminating DNAs in molecular phylogenetic studies, particularly when using "universal" amplification primers.
authors.affiliation: Department of Botany, Iowa State University, Ames, Iowa, 50011, USA.
authors.forename: W
authors.initials: W
authors.lastname: Zhang
authors.forename: J F
authors.initials: JF
authors.lastname: Wendel
authors.forename: L G
authors.initials: LG
authors.lastname: Clark
mesh_terms: D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics* / Q000302:isolation & purification*; D018744:DNA, Plant / Q000235:genetics* / Q000302:isolation & purification*; D004275:DNA, Ribosomal / Q000235:genetics* / Q000302:isolation & purification*; D005658:Fungi / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D008855:Microscopy, Electron, Scanning; D008969:Molecular Sequence Data; D010802:Phylogeny; D010944:Plants / Q000145:classification / Q000235:genetics* / Q000382:microbiology*; D016133:Polymerase Chain Reaction; D012340:RNA, Ribosomal, 5.8S / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D018744:DNA, Plant; D004275:DNA, Ribosomal; D012340:RNA, Ribosomal, 5.8S