A new fungal large subunit ribosomal RNA primer for high-throughput sequencing surveys

被引:39
作者
Mueller, Rebecca C. [1 ]
Gallegos-Graves, La Verne [1 ]
Kuske, Cheryl R. [1 ]
机构
[1] Los Alamos Natl Lab, Biosci Div, M888 HRL, Los Alamos, NM 87545 USA
关键词
Illumina sequencing; large subunit ribosomal RNA gene; fungal community composition; phylogenetic community measures; contrived community analysis; INTERNAL TRANSCRIBED SPACER; MAXIMUM-LIKELIHOOD; COMMUNITIES; ACCURATE; CLASSIFICATION; EVOLUTION; RESPONSES; REGION; TREES; PLANT;
D O I
10.1093/femsec/fiv153
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
The inclusion of phylogenetic metrics in community ecology has provided insights into important ecological processes, particularly when combined with high-throughput sequencing methods; however, these approaches have not been widely used in studies of fungal communities relative to other microbial groups. Two obstacles have been considered: (1) the internal transcribed spacer (ITS) region has limited utility for constructing phylogenies and (2) most PCR primers that target the large subunit (LSU) ribosomal unit generate amplicons that exceed current limits of high-throughput sequencing platforms. We designed and tested a PCR primer (LR22R) to target approximately 300-400 bp region of the D2 hypervariable region of the fungal LSU for use with the Illumina MiSeq platform. Both in silico and empirical analyses showed that the LR22R-LR3 pair captured a broad range of fungal taxonomic groups with a small fraction of non-fungal groups. Phylogenetic placement of publically available LSU D2 sequences showed broad agreement with taxonomic classification. Comparisons of the LSU D2 and the ITS2 ribosomal regions from environmental samples and known communities showed similar discriminatory abilities of the two primer sets. Together, these findings show that the LR22R-LR3 primer pair has utility for phylogenetic analyses of fungal communities using high-throughput sequencing methods.
引用
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页码:1 / 11
页数:11
相关论文
共 49 条
[21]   Reconstructing the early evolution of Fungi using a six-gene phylogeny [J].
James, Timothy Y. ;
Kauff, Frank ;
Schoch, Conrad L. ;
Matheny, P. Brandon ;
Hofstetter, Valerie ;
Cox, Cymon J. ;
Celio, Gail ;
Gueidan, Cecile ;
Fraker, Emily ;
Miadlikowska, Jolanta ;
Lumbsch, H. Thorsten ;
Rauhut, Alexandra ;
Reeb, Valerie ;
Arnold, A. Elizabeth ;
Amtoft, Anja ;
Stajich, Jason E. ;
Hosaka, Kentaro ;
Sung, Gi-Ho ;
Johnson, Desiree ;
O'Rourke, Ben ;
Crockett, Michael ;
Binder, Manfred ;
Curtis, Judd M. ;
Slot, Jason C. ;
Wang, Zheng ;
Wilson, Andrew W. ;
Schuessler, Arthur ;
Longcore, Joyce E. ;
O'Donnell, Kerry ;
Mozley-Standridge, Sharon ;
Porter, David ;
Letcher, Peter M. ;
Powell, Martha J. ;
Taylor, John W. ;
White, Merlin M. ;
Griffith, Gareth W. ;
Davies, David R. ;
Humber, Richard A. ;
Morton, Joseph B. ;
Sugiyama, Junta ;
Rossman, Amy Y. ;
Rogers, Jack D. ;
Pfister, Don H. ;
Hewitt, David ;
Hansen, Karen ;
Hambleton, Sarah ;
Shoemaker, Robert A. ;
Kohlmeyer, Jan ;
Volkmann-Kohlmeyer, Brigitte ;
Spotts, Robert A. .
NATURE, 2006, 443 (7113) :818-822
[22]   MAFFT Multiple Sequence Alignment Software Version 7: Improvements in Performance and Usability [J].
Katoh, Kazutaka ;
Standley, Daron M. .
MOLECULAR BIOLOGY AND EVOLUTION, 2013, 30 (04) :772-780
[23]   Plant traits and taxonomy drive host associations in tropical phyllosphere fungal communities [J].
Kembel, Steven W. ;
Mueller, Rebecca C. .
BOTANY, 2014, 92 (04) :303-311
[24]   Incorporating 16S Gene Copy Number Information Improves Estimates of Microbial Diversity and Abundance [J].
Kembel, Steven W. ;
Wu, Martin ;
Eisen, Jonathan A. ;
Green, Jessica L. .
PLOS COMPUTATIONAL BIOLOGY, 2012, 8 (10)
[25]   Global diversity and distribution of arbuscular mycorrhizal fungi [J].
Kivlin, Stephanie N. ;
Hawkes, Christine V. ;
Treseder, Kathleen K. .
SOIL BIOLOGY & BIOCHEMISTRY, 2011, 43 (11) :2294-2303
[26]   Development of a Dual-Index Sequencing Strategy and Curation Pipeline for Analyzing Amplicon Sequence Data on the MiSeq Illumina Sequencing Platform [J].
Kozich, James J. ;
Westcott, Sarah L. ;
Baxter, Nielson T. ;
Highlander, Sarah K. ;
Schloss, Patrick D. .
APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2013, 79 (17) :5112-5120
[27]  
Letunic I, 2011, NUCLEIC ACIDS RES, V39, pW475, DOI [10.1093/nar/gkr201, 10.1093/nar/gkr931]
[28]   Employing 454 amplicon pyrosequencing to reveal intragenomic divergence in the internal transcribed spacer rDNA region in fungi [J].
Lindner, Daniel L. ;
Carlsen, Tor ;
Nilsson, R. Henrik ;
Davey, Marie ;
Schumacher, Trond ;
Kauserud, Havard .
ECOLOGY AND EVOLUTION, 2013, 3 (06) :1751-1764
[29]   RAxML and FastTree: Comparing Two Methods for Large-Scale Maximum Likelihood Phylogeny Estimation [J].
Liu, Kevin ;
Linder, C. Randal ;
Warnow, Tandy .
PLOS ONE, 2011, 6 (11)
[30]   Accurate, Rapid Taxonomic Classification of Fungal Large-Subunit rRNA Genes [J].
Liu, Kuan-Liang ;
Porras-Alfaro, Andrea ;
Kuske, Cheryl R. ;
Eichorst, Stephanie A. ;
Xie, Gary .
APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2012, 78 (05) :1523-1533