Project description:We have been studying intersex in male rainbow darter (Etheostoma caeruleum) associated with exposure to sewage effluents. To understand changes in the gene transcriptome associated with intersex it was necessary to have a better understanding of normal annual changes in the transcriptome. The goal of this research is to identify patterns of gene expression associated with the different stages of gonad development during the annual cycle. The studies of molecular pathways involve in ovarian or testis development has been poorly studied. While most studies focus on female ovarian changes, there is a gap in understanding testis development. A customized second generation microarray for rainbow darter (8x15k) was used to identify patterns of gene expression -in terms of mRNA abundance- in male rainbow darter gonads during an annual cycle. Rainbow darter males were collected on field work surveys in May (spawning), August (post-spawning), and October (recrudescence) 2011, and January (developing) and March (pre-spawning) 2012, using a back pack electrofisher from a clean area at the Grand River, ON, Canada.
Project description:We have been studying intersex in male rainbow darter (Etheostoma caeruleum) associated with exposure to sewage effluents. To understand changes in the gene transcriptome associated with intersex it was necessary to have a better understanding of normal annual changes in the transcriptome. The goal of this research is to identify patterns of gene expression associated with the different stages of gonad development during the annual cycle. The studies of molecular pathways involve in ovarian or testis development has been poorly studied. While most studies focus on female ovarian changes, there is a gap in understanding testis development. A customized second generation microarray for rainbow darter (8x15k) was used to identify patterns of gene expression -in terms of mRNA abundance- in male rainbow darter gonads during an annual cycle.
Project description:In this study, adult male rainbow darter (Etheostoma caeruleum) were collected from nine sites located at different distances from two wastewater treatment plants (WWTPs) in the Grand River, Ontario, Canada. The goal of this project was to see if the liver transcriptome of fish residing downstream of the two WWTPs differs from the non-exposed fish. Further, this research was also addressing the affects of differnent reference site selection on gene expression patterns, and their subsequent interpretation.The RNA from male rainbow darter was exctracted from liver tissue and used for further gene expression analysis.
Project description:The utility of RADseq in an experimental setting is also demonstrated, based on our chasacterisation of an APOBEC mutation signature in an APOBEC3A transfected mouse cell line. 0D5 cells, derived from SSM3 cells, were co-transfected with a mixture containing pcDNA3.1 vectors expressing either APOBEC3A or APOBEC3B (kindly donated by Vincent Caval), pcDNA3.1 construct expressing deaminase null APOBEC3A linked to a uracil deglycosylase construct and a plasmid encoding mutant GFP and WT mCherry that is a reporter for APOBEC mutagenesis. Cells were grown, and gDNA extracted, prior to preparation of RADseq libraries using a PstI- MspI double-digest. Libraries underwent a Pippin Prep to select fragments in the size range of 220-520 bp (genomic sequence plus 148 bp of adapters). Single-end sequencing (1x101bp) was performed on an Illumina NovaSeq6000 utilizing v1.5 chemistry. Reads were aligned to mm10 using bwa mem and variants called using the GATK4 pipeline.
Project description:In this study, adult male rainbow darter were collected from three sites in the Grand River in the fall season for four consecutive years (2011-2014), from 3 sites: a reference site (US2) located in the non-urbanized area, and two exposure sites (DSW and DSK1) located dowstrem of two secondary wastewater treatment palnts (WWTP) (Waterloo and Kitchener). Kitchener WWTP has been undergone infrastructure upgrades that were initiated in the summer of 2012. The goal of this project was to see whether the implemented upgrades can be reflected in the liver transcriptome response indicative of recovery.The RNA from male rainbow darter was exctracted from liver tissue and used for further gene expression analysis.
2017-02-21 | GSE95083 | GEO
Project description:ddRADseq Relict Darter (E. chienense) and Clarks Darter (E. cf. oophylax)
Project description:Rainbow darter (Etheostoma caeruleum) are a small benthic fish found in North America. This species is sensitive to sewage effluent in the environment, showing the presence of intersex in up to 80% of males in near-field areas in the Grand River, ON. To learn more about the molecular events associated with intersex, we developed a customized oligonucleotide microarray (4x180K) with next generation sequencing (454 Roche) to characterize molecular responses in the gonad. Transcriptomics was performed on both males and females from both a reference site and a polluted site. Males with and without intersex from the polluted site were compared to the control males. Rainbow darter were sampled from from the Grand River in May 2011. Fish were selected according to the location, gonad maturity, and intersex index. Reference fish were taken from the upstream to the urban area; exposed fish were taken from downstream of from Kitchener MWWE treatment plant.