Project description:Today, nanoparticles are used in various commercial products. One of the most common nanoparticles is titanium dioxide (TiO2). It has a catalytic activity and UV absorption (λ<400 nm), and it generates reactive oxygen species (ROS). The catalytic activity of TiO2 nanoparticle is capable of killing a wide range of microorganisms. In the environment, nanoparticles form structures consisting of primary particles, and their aggregates and agglomerates. These compounds are defined NOAA (nano-objects, and their aggregates and agglomerates greater than 100 nm). The unique properties of TiO2 nanoparticles can be maintained in the environment, thus, the growing use of TiO2 nanoparticles is raising concerns about the environmental risks. The assessment of biological and ecological effects of TiO2-NOAA is necessary. In our previous study, we assessed the effect of TiO2-NOAA on microbes by using Saccharomyces cerevisiae and Escherichia coli. It was shown that TiO2-NOAA decomposed methylene blue under UV irradiation. It suggested that TiO2-NOAA generated ROS under UV irradiation. However, TiO2-NOAA did not show growth inhibition in minimal agar medium under UV irradiation. By adding TiO2-NOAA in medium, colony formation was observed with UV intensity that inactivates microbes. Moreover, TiO2-NOAA adsorbed microbes. These results suggested that the amount of ROS generated by TiO2-NOAA was not enough to inactivate microbes, and TiO2-NOAA might protect microbes from UV. In this study, we assessed the effect of TiO2-NOAA in more detail by using S. cerevisiae. We used DNA microarray analysis for qualitative assessment. Further, we carried out quantitative assessment by using Real Time RT-PCR method for characteristic genes in DNA microarray analysis. To compare yeast cells in various conditions, six kinds of treatment conditions were prepared (Condition 1. adsorbed fraction to TiO2-NOAA under UV, 2. non-adsorbed fraction to TiO2-NOAA under UV, 3. adsorbed fraction to TiO2-NOAA without UV, 4. non-adsorbed fraction to TiO2-NOAA without UV, 5. irradiated UV and 6. negative control). From the result of DNA microarray analysis, the most number of genes was altered in Condition 1, followed by Condition 3 and 5. The genes related to oxidative stress, and the genes related to synthesis of trehalose and glycogen were significantly up-regulated of yeast cells in Condition 1 and 5, and Condition 1 and 3, respectively. These results suggest that yeast cells suffer oxidative stress by TiO2-NOAA under UV, and they also suffer membrane damage by TiO2-NOAA itself, as a result, they reserve energy sources. From the result of Real Time RT-PCR, genes related to oxidative stress (GRE2, SOD2) were up-regulated in Condition 1 and 3, however, these expression levels in each condition were not significant. And genes related to synthesis of trehalose and glycogen (GSY1, TPS2) were up-regulated in Condition 1 and 3. These results suggest that oxidative stress is caused not by TiO2-NOAA but by UV. It is also suggested that yeast cells were damaged at their membranes by TiO2-NOAA, as a result, genes related to synthesis of trehalose and glycogen were up-regulated. Thus, we suggest that the effect of TiO2-NOAA on yeast cells under UV irradiation is greater due to TiO2-NOAA itself than due to ROS generated by TiO2-NOAA.

Project description:To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild.

Project description:To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. The expanded genome proxy array was designed as in (Rich et al., 2008). Briefly, each genotype was targeted using suites of ~20 70-mer oligonucleotide probes designed using the program ArrayOligoSelector (Zhu et al., 2003). Probes had approximately the same %GC (40%) and were distributed across the target genome or genome fragment, with no more than one probe per gene and avoiding 16S and 23S rRNA genes. The array included positive and negative control probes designed using the same method, to Halobacterium salinarum NRC-1 and a random genome sequence, respectively. The expanded array had a broader scope than the prototype of Rich et al., 2008 (268 target genotypes, as opposed to the prototype’s 14) and included a co-spot oligo for spot alignment and gridding purposes (using the “alien” oligo sequence of (Urisman et al., 2005). The targets were selected from fully-sequenced marine microbial genomes, publicly-available marine-derived BAC and fosmid clone sequences, and fully-sequenced clones from the lab’s Monterey Bay and Hawaii environmental BAC- and fosmid-based genomic libraries. Targeted genotypes are detailed in Table S1, summarized in Table S2, and presented in a schematic phylogenetic overview in Fig. 1. Previously-unpublished sequences used for array design were submitted to Genbank under accession numbers GU474833-GU474949. This submission presents the analysis of 57 samples, each hybridized to >= 3 arrays. In addition to negative control probes, for each hyrbdization date and run a negative control array was hyrbdized for which the input DNA was TE, taken through the entire aplification, labeling, and hybridization process.

			Action	DRS
	SRR14163830_1.fastq.gz	Fastqsanger.gz
	SRR14163830_2.fastq.gz	Fastqsanger.gz
	SRR14433900_1.fastq.gz	Fastqsanger.gz
	SRR14433900_2.fastq.gz	Fastqsanger.gz
	SRR14433901_1.fastq.gz	Fastqsanger.gz

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