Research Accomplishment Reports 2011

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Advanced Genetic Technologies, KY

C.L. Schardl
Department of Plant Pathology

 

Non-Technical Summary

DNA sequencing technologies and associated methods for analyzing DNA sequences have become so advanced and affordable that rapid sequencing of the entire genetic constitution (whole genome sequencing) of microbial species is now routine. Application of whole genome sequencing to parasites and symbionts of plants and animals allows deep insight into their interactions with hosts, and can be used to address productivity and diseases of crops and livestock. Such insights require facile analysis of the very large datasets generated by whole genome sequencing.

This project will use, as a test case, a group of fungi that range from parasitic to beneficial in their interactions with common forage grasses and wild grass relatives. Whole genomes will be sequenced from 14 of these fungi, sets of closely homologous genes will be identified and their sequences will be compared in order to discern the evolutionary relationships of those genes. The results will provide insight into the underlying genetics of plant pathogenesis as well as beneficial symbiosis.

These studies will require streamlining and standardizing computer analysis of the large volume of data obtained by whole genome sequencing. Therefore, a central activity will be to refine computer methods and algorithms for acquisition, collation, management and analysis of these data, thereby greatly enhancing the bioinformatic capabilities at the University of Kentucky.

2011 Project Description

Genome sequences were determined for twelve genomes from fungi in genera Aciculosporium, Claviceps, Colletotrichum (i.e., Glomerella), Epichloe, Neotyphodium and Periglandula. These fungi were chosen for the potential to relate their gene contents to their diverse interactions with grass species, ranging from biotrophic (colonizing only living plant tissue) to hemibiotrophic (colonizing living tissue, then killing and further colonizing host cells), and from mutualistic symbionts to pathogens.

Considerable progress was made in establishing computational pipelines for gene annotation, comparative genomics and phylogenetics. Gene models were constructed based on combinations of ab initio gene predictions and biological data in the form of messenger RNA (mRNA) sequences. Gene functions are assigned based on results of homology searches with Blast2GO, and hidden markov model (HMM) searches of the sequences for signatures that identify protein and enzyme families.

The comparative genomics pipeline employed two computer algorithms run in series to identify sets of orthologous genes between genomes, and to align the sequences of orthologs to facilitate phylogenetic analysis on a genome-wide scale.

This project served as a key training ground for staff and students in high-throughput genome (DNA) and transcriptome (RNA) sequencing and annotation. A bioinformatician attended the blast2GO workshop and undertook bioinformatic analysis of data generated in the DNA sequencing core facility.

2011 Impact

Genome sequences, together with ortholog groups and alignments of protein sequences inferred from the annotated genomes, have been published to a web site, www.endophyte.uky.edu, and also have been uploaded to the Genbank database at the National Center for Biotechnology Information (NCBI).

2011 Publications

Chambers EW, Hapairai L, Peel BA, Bossin H, Dobson SL (2011) Male Mating Competitiveness of a Wolbachia-introgressed Aedes polynesiensis strain under semi-field conditions. PLoS Neglected Tropical Diseases 5, doi: 10.1371/journal.pntd.0001271

D'Angelo E, Nunez A (2010) Effect of environmental conditions on polychlorinated biphenyl transformations and bacterial communities in a river sediment. Journal of Soils and Sediments 10: 1186-1199. doi: 10.1007/s11368-010-0218-2

Gao QM, Venugopal S, Navarre D, Kachroo A (2011) Low Oleic Acid-Derived Repression of jasmonic acid-inducible defense responses requires the WRKY50 and WRKY51 proteins. Plant Physiology 155: 464-476. doi: 10.1104/pp.110.166876

Huang TS, Nagy PD (2011) Direct inhibition of Tombusvirus plus-strand RNA synthesis by a dominant negative mutant of a host metabolic enzyme, glyceraldehyde-3-phosphate dehydrogenase, in yeast and plants. Journal of Virology 85: 9090-9102. doi: 10.1128/jvi.00666-11

Ji LJ, Liu XG, Yan J, Wang WM, Yumul RE, Kim YJ, Dinh TT, Liu J, Cui X, Zheng BL, Agarwal M, Liu CY, Cao XF, Tang GL, Chen XM (2011) ARGONAUTE10 and ARGONAUTE1 regulate the termination of floral stem cells through two MicroRNAs in Arabidopsis. PLoS Genetics 7doi: 10.1371/journal.pgen.1001358

Nagy PD, Wang RY, Pogany J, Hafren A, Makinen K (2011) Emerging picture of host chaperone and cyclophilin roles in RNA virus replication. Virology 411: 374-382. doi: 10.1016/j.virol.2010.12.061

Sharanowski BJ, Dowling APG, Sharkey MJ (2011) Molecular phylogenetics of Braconidae (Hymenoptera: Ichneumonoidea), based on multiple nuclear genes, and implications for classification. Systematic Entomology 36: 549-572. doi: 10.1111/j.1365-3113.2011.00580.x

Stork J, Kovalev N, Sasvari Z, Nagy PD (2011) RNA chaperone activity of the tombusviral p33 replication protein facilitates initiation of RNA synthesis by the viral RdRp in vitro. Virology 409: 338-347. doi: 10.1016/j.virol.2010.10.015

Venard CMP, Crain PR, Dobson SL (2011) SYTO11 staining vs FISH staining: a comparison of two methods to stain Wolbachia pipientis in cell cultures. Letters in Applied Microbiology 52: 168-176. doi: 10.1111/j.1472-765X.2010.02986.x