Project description:We used custom Nimblegen microarrays representing whole-larval transcriptomes for two species (Erynnis propertius [this submission] and Papilio zelicaon [submitted seperately]) to assess gene expression differences affecting tolerance to climatic regimes. Many individuals were sourced from populations from the northern periphery and center of the species' (shared) range; these were each divided into groups treated under peripheral and central climate regimes, resulting in 4 experimental groups for each species (Peripheral Source, Peripheral treatment; Peripheral Source, Central Treatment; Central Source, Peripheral Treatment; Central Source, Central Treatment). Using technical microarray replicates allowed us to use ANOVA to identify genes whose expression may underlie local adaptation to climate (i.e., those showing an interaction term between source and population). Abstract: Population differences may determine geographic range shifts and adaptive evolution under climate change. Local adaptation in peripheral populations could preclude or slow range expansions, and populations with different genetic make-up could have distinct trajectories that produce complex spatial patterns of population change. To investigate the genetic extent of local responses to climate change, we exposed poleward-periphery and central populations of two Lepidoptera to reciprocal, common-garden climatic conditions and compared whole-transcriptome expression. We found significant expression differences between populations in both species. In addition, several hundred genes including genes involved in energy metabolism and oxidative stress responded in a localized fashion in the species that exhibits greater population structure and local adaptation. Expression levels of these genes are most divergent in the same environment in which we previously detected phenotypic divergence in metabolism. By contrast, we found no localized genes in the species with higher gene flow, reflecting the lack of previously observed local adaptation. These results suggest that population differences do not generalize easily, even for related species living in the same climate, but some taxa deserve population-level consideration when predicting the effects of climate change.

Project description:We used custom Nimblegen microarrays representing whole-larval transcriptomes for two species (Erynnis propertius [this submission] and Papilio zelicaon [submitted seperately]) to assess gene expression differences affecting tolerance to climatic regimes. Many individuals were sourced from populations from the northern periphery and center of the species' (shared) range; these were each divided into groups treated under peripheral and central climate regimes, resulting in 4 experimental groups for each species (Peripheral Source, Peripheral treatment; Peripheral Source, Central Treatment; Central Source, Peripheral Treatment; Central Source, Central Treatment). Using technical microarray replicates allowed us to use ANOVA to identify genes whose expression may underlie local adaptation to climate (i.e., those showing an interaction term between source and population). Abstract: Population differences may determine geographic range shifts and adaptive evolution under climate change. Local adaptation in peripheral populations could preclude or slow range expansions, and populations with different genetic make-up could have distinct trajectories that produce complex spatial patterns of population change. To investigate the genetic extent of local responses to climate change, we exposed poleward-periphery and central populations of two Lepidoptera to reciprocal, common-garden climatic conditions and compared whole-transcriptome expression. We found significant expression differences between populations in both species. In addition, several hundred genes including genes involved in energy metabolism and oxidative stress responded in a localized fashion in the species that exhibits greater population structure and local adaptation. Expression levels of these genes are most divergent in the same environment in which we previously detected phenotypic divergence in metabolism. By contrast, we found no localized genes in the species with higher gene flow, reflecting the lack of previously observed local adaptation. These results suggest that population differences do not generalize easily, even for related species living in the same climate, but some taxa deserve population-level consideration when predicting the effects of climate change. Previously we sequenced and assembled whole larval transcriptome ESTs sourced from pooled central-population individuals subjected to environmental stressors (see O'Neil et al., 2008). From these assemblies custom Nimblegen microarrays were designed (Nimblegen, Inc.), representing 34,609 putative gene sequences for E. propertius (this submission) and 25,735 putative gene sequences for P. zelicaon (submitted seperately). Probe designs sought 5 representative 60mer probes for E.propertius and 4 representative probes for P. zelicaon. Messenger RNA was was sampled from multiple individuals of each experimental group and pooled before being converted to cDNA and hybridized to technical replicate microarrays. Three technical replicates for each experimental group were used, for a total of 12 microarrays (per species). Microarray data were log2 transformed and quintile-normalized (Bolstad et al. 2003) on a per-species basis.

Project description:BackgroundSeveral recent studies have demonstrated the use of Roche 454 sequencing technology for de novo transcriptome analysis. Low error rates and high coverage also allow for effective SNP discovery and genetic diversity estimates. However, genetically diverse datasets, such as those sourced from natural populations, pose challenges for assembly programs and subsequent analysis. Further, estimating the effectiveness of transcript discovery using Roche 454 transcriptome data is still a difficult task.ResultsUsing the Roche 454 FLX Titanium platform, we sequenced and assembled larval transcriptomes for two butterfly species: the Propertius duskywing, Erynnis propertius (Lepidoptera: Hesperiidae) and the Anise swallowtail, Papilio zelicaon (Lepidoptera: Papilionidae). The Expressed Sequence Tags (ESTs) generated represent a diverse sample drawn from multiple populations, developmental stages, and stress treatments. Despite this diversity, > 95% of the ESTs assembled into long (> 714 bp on average) and highly covered (> 9.6x on average) contigs. To estimate the effectiveness of transcript discovery, we compared the number of bases in the hit region of unigenes (contigs and singletons) to the length of the best match silkworm (Bombyx mori) protein--this "ortholog hit ratio" gives a close estimate on the amount of the transcript discovered relative to a model lepidopteran genome. For each species, we tested two assembly programs and two parameter sets; although CAP3 is commonly used for such data, the assemblies produced by Celera Assembler with modified parameters were chosen over those produced by CAP3 based on contig and singleton counts as well as ortholog hit ratio analysis. In the final assemblies, 1,413 E. propertius and 1,940 P. zelicaon unigenes had a ratio > 0.8; 2,866 E. propertius and 4,015 P. zelicaon unigenes had a ratio > 0.5.ConclusionsUltimately, these assemblies and SNP data will be used to generate microarrays for ecoinformatics examining climate change tolerance of different natural populations. These studies will benefit from high quality assemblies with few singletons (less than 26% of bases for each assembled transcriptome are present in unassembled singleton ESTs) and effective transcript discovery (over 6,500 of our putative orthologs cover at least 50% of the corresponding model silkworm gene).

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Erynnis propertius

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