市町村合併における財政指標の利用に関する基礎的研究

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RNA-Seq reveals virus–virus and virus–plant interactions in nature

As research on plant viruses has focused mainly on crop diseases, little is known about these viruses in natural environments. To understand the ecology of viruses in natural systems, comprehensive information on virus–virus and virus–host interactions is required. We applied RNA-Seq to plants from a natural population of Arabidopsis halleri subsp. gemmifera to simultaneously determine the presence/absence of all sequence-reported viruses, identify novel viruses and quantify the host transcriptome. By introducing the criteria of read number and genome coverage, we detected infections by Turnip mosaic virus (TuMV), Cucumber mosaic virus and Brassica yellows virus. Active TuMV replication was observed by ultramicroscopy. De novo assembly further identified a novel partitivirus, Arabidopsis halleri partitivirus 1. Interestingly, virus reads reached a maximum level that was equivalent to that of the host's total mRNA, although asymptomatic infection was common. AhgAGO2, a key gene in host defence systems, was upregulated in TuMV-infected plants. Multiple infection was frequent in TuMV-infected leaves, suggesting that TuMV facilitates multiple infection, probably by suppressing host RNA silencing. Revealing hidden plant–virus interactions in nature can enhance our understanding of biological interactions and may have agricultural applications

Neighbor GWAS: incorporating neighbor genotypic identity into genome-wide association studies of field herbivory

An increasing number of field studies have shown that the phenotype of an individual plant depends not only on its genotype but also on those of neighboring plants; however, this fact is not taken into consideration in genome-wide association studies (GWAS). Based on the Ising model of ferromagnetism, we incorporated neighbor genotypic identity into a regression model, named "Neighbor GWAS". Our simulations showed that the effective range of neighbor effects could be estimated using an observed phenotype when the proportion of phenotypic variation explained (PVE) by neighbor effects peaked. The spatial scale of the first nearest neighbors gave the maximum power to detect the causal variants responsible for neighbor effects, unless their effective range was too broad. However, if the effective range of the neighbor effects was broad and minor allele frequencies were low, there was collinearity between the self and neighbor effects. To suppress the false positive detection of neighbor effects, the fixed effect and variance components involved in the neighbor effects should be tested in comparison with a standard GWAS model. We applied neighbor GWAS to field herbivory data from 199 accessions of Arabidopsis thaliana and found that neighbor effects explained 8% more of the PVE of the observed damage than standard GWAS. The neighbor GWAS method provides a novel tool that could facilitate the analysis of complex traits in spatially structured environments and is available as an R package at CRAN ( https://cran.rproject.org/package=rNeighborGWAS )

An efficient early-pooling protocol for environmental DNA metabarcoding

Environmental DNA (eDNA) metabarcoding, a method that applies high-throughput sequencing and universal primer sets to eDNA analysis, has been a promising approach for efficient, comprehensive biodiversity monitoring. However, significant money-, labor-, and time-costs are still required for performing eDNA metabarcoding. In this study, we assessed the performance of an “early-pooling” protocol (a protocol based on 1st PCR tagging) to reduce the experimental costs of library preparation for eDNA metabarcoding. Specifically, we performed three experiments to investigate the effects of 1st PCR-tagging and 2nd PCR-indexing protocols on the community composition revealed by eDNA metabarcoding, the effects of post-1st PCR exonuclease purification on tag jumping (corresponds to index hopping in 2nd PCR indexing), and the effects of the number of PCR replicates and the eDNA template volume on the number of detected OTUs. Analyses of 204 eDNA libraries from three natural aquatic ecosystems and one mock eDNA sample showed that (i) 1st PCR tagging does not cause clear biases in the outcomes of eDNA metabarcoding, (ii) post-1st PCR exonuclease purification reduces the risk of tag jumping, and (iii) increasing the eDNA template volume may increase the number of detected OTUs and reduce variations in the detected community compositions, similar to increasing the number of 1st PCR replicates. Our results show that an early-pooling protocol with post-1st PCR exonuclease purification and an increased amount of the DNA template reduces the risk of tag jumping, the costs for consumables and reagents (except for many tagged 1st PCR primers), and the handling time in library preparation, and produces similar results to a 2nd PCR-indexing protocol. Therefore, once a target metabarcoding region is selected and a set of tagged-1st PCR primers is prepared, the early-pooling protocol provides a cost, labor, and time-efficient approach for processing a large number of samples

Genetic population structure of the precious coral Corallium japonicum in the Northwest Pacific

Population sizes of the Japanese red coral Corallium japonicum have been severely affected by poaching and overfishing. Although genetic structure and connectivity patterns are considered important parameters for conservation strategies, there are few studies focusing on the population genetics of C. japonicum in the Northwest Pacific. We examined the genetic population structure of C. japonicum, in the Northwest Pacific. We used restriction-site-associated DNA sequencing (RAD-seq), which can be used to identify genome-wide single-nucleotide polymorphism (SNPs), to reveal detailed within-species genetic variations. Using the variable SNP loci identified from this analysis, we successfully evaluated the population-level genetic diversity and patterns of gene flow among multiple populations of C. japonicum around Japan. The results of genetic analysis basically showed that gene flow is widely maintained in the geographic range examined in this study, but the analysis in combination with larval dispersal simulations revealed several populations that were genetically distinct from the other populations, suggesting geographically limited gene flows. The information obtained from this study will be useful for the design of effective management schemes for C. japonicum, which is under threat from overfishing

Detection of diurnal variation of tomato transcriptome through the molecular timetable method in a sunlight-type plant factory

The timing of measurement during plant growth is important because many genes are expressed periodically and orchestrate physiological events. Their periodicity is generated by environmental fluctuations as external factors and the circadian clock as the internal factor. The circadian clock orchestrates physiological events such as photosynthesis or flowering and it enables enhanced growth and herbivory resistance. These characteristics have possible applications for agriculture. In this study, we demonstrated the diurnal variation of the transcriptome in tomato (Solanum lycopersicum) leaves through molecular timetable method in a sunlight-type plant factory. Molecular timetable methods have been developed to detect periodic genes and estimate individual internal body time from these expression profiles in mammals. We sampled tomato leaves every 2 h for 2 days and acquired time-course transcriptome data by RNA-Seq. Many genes were expressed periodically and these expressions were stable across the 1st and 2nd days of measurement. We selected 143 time-indicating genes whose expression indicated periodically, and estimated internal time in the plant from these expression profiles. The estimated internal time was generally the same as the external environment time; however, there was a difference of more than 1 h between the two for some sampling points. Furthermore, the stress-responsive genes also showed weakly periodic expression, implying that they were usually expressed periodically, regulated by light–dark cycles as an external factor or the circadian clock as the internal factor, and could be particularly expressed when the plant experiences some specific stress under agricultural situations. This study suggests that circadian clock mediate the optimization for fluctuating environments in the field and it has possibilities to enhance resistibility to stress and floral induction by controlling circadian clock through light supplement and temperature control

自治体組織の問題点と小樽市における電子的住民投票システムの活用効果について

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