Research Accomplishment Reports 2011

Advanced Genetic Technologies, KY

C.L. Schardl
Department of Plant Pathology

 

Non-Technical Summary

The genome of an organism represents the total of all genes, which are encoded in the base sequences of DNA of that organism. In order for a gene to be expressed, it must first be transcribed into RNA molecules, each similar in sequence to a DNA segment. Then, the RNAs are spliced and processed into mRNAs, which are then translated to give functional proteins. The entire set of mRNAs in a cell, tissue or organism is called its transcriptome. Analysis of whole-transcriptome sequence is key to understanding how genomes direct the organism's form, development, behavior, and responses to the environment. New technologies such as pyrosequencing have greatly increased DNA sequencing rates and reduced costs. In this study, several strategies will be tested for pyrosequencing of transcriptomes. Once these protocols are refined, they will be applied to various organisms of relevance to agricultural research, including plants, insects, and various pathogens and parasites. The results will serve as preliminary data for competitively funded research programs.

2011 Project Description

Several RNA (transcriptome) sequencing projects were undertaken on plants, insects, fungi, and symbiotic microbes responsible for cellulose digestion in wood roaches and termites. Two methods were compared: the Roche/454 rapid cDNA sequencing system, and the Matz protocol (Meyer E, Aglyamova G, Wang S, Buchanan-Carter J, Abrego D, Colbourne J, Willis B, Matz M (2009) Sequencing and de novo analysis of a coral larval transcriptome using 454 GSFlx. BMC Genomics 10: 219.)

Also, assembly programs for transcriptome sequence were compared, including Velvet, CLC Genome Workbench, and MapSplice.

Finally, the bioinformatition assigned to the AGTC received training in the Blast2GO genome and transcriptome annotation system.

2011 Impact

The Roche/454 rapid cDNA sequencing system proved to be feasible for most applications. The Matz protocol was considerably more expensive and time-consuming, but provided more specific information that enhanced accuracy of gene models generated from the data. Strengths and weaknesses of the various software packages were also identified. The results of these studies provide a basis for standard operating procedures and recommended protocols to be used by the AGTC core facility

2011 Publications

Brelsfoard CL, Dobson SL (2011) Wolbachia effects on host fitness and the influence of male aging on cytoplasmic incompatibility in Aedes polynesiensis (Diptera: Culicidae). Journal of Medical Entomology 48: 1008-1015. doi: 10.1603/me10202

Fu YQ, Gavotte L, Mercer DR, Dobson SL (2010) Artificial triple Wolbachia infection in Aedes albopictus yields a new pattern of unidirectional cytoplasmic incompatibility. Applied and Environmental Microbiology 76: 5887-5891. doi: 10.1128/aem.00218-10

Heraty J, Ronquist F, Carpenter JM, Hawks D, Schulmeister S, Dowling AP, Murray D, Munro J, Wheeler WC, Schiff N, Sharkey M (2011) Evolution of the hymenopteran megaradiation. Molecular Phylogenetics and Evolution 60: 73-88. doi: 10.1016/j.ympev.2011.04.003

Li ZH, Nagy PD (2011) Diverse roles of host RNA-binding proteins in RNA virus replication. RNA Biology 8: 305-315. doi: 10.4161/rna.8.2.15391

Sharma M, Sasvari Z, Nagy PD (2011) Inhibition of phospholipid biosynthesis decreases the activity of the Tombusvirus replicase and alters the subcellular localization of replication proteins. Virology 415: 141-152. doi: 10.1016/j.virol.2011.04.008

Steiner U, Leibner S, Schardl CL, Leuchtmann A, Leistner E (2011) Periglandula, a new fungal genus within the Clavicipitaceae and its association with Convolvulaceae. Mycologia 103: 1133-1145. doi: 10.3852/11-031

Tsyusko O, Glenn T, Yi Y, Joice G, Jones K, Aizawa K, Coughlin D, Zimbrick J, Hinton T (2011) Differential genetic responses to ionizing irradiation in individual families of Japanese medaka, Oryzias latipes. Mutation Research-Genetic Toxicology and Environmental Mutagenesis 718: 18-23. doi: 10.1016/j.mrgentox.2010.11.001

Wang SH, Kurepa J, Hashimoto T, Smalle JA (2011) Salt stress-induced disassembly of Arabidopsis cortical microtubule arrays involves 26S proteasome-dependent degradation of SPIRAL1. Plant Cell 23: 3412-3427. doi: 10.1105/tpc.111.089920

Zhu SF, Jeong RD, Venugopal SC, Lapchyk L, Navarre D, Kachroo A, Kachroo P (2011) SAG101 forms a ternary complex with EDS1 and PAD4 and is required for resistance signaling against turnip crinkle virus. PLoS Pathogens 7doi: 10.1371/journal.ppat.1002318