Project description:Nuclear and mitochondrial organelles must maintain a communication system. Loci on the mitochondrial genome were recently reported to interact with nuclear loci. To determine whether this is part of a DNA based communication system we used genome conformation capture to map the global network of DNA-DNA interactions between the mitochondrial and nuclear genomes (Mito-nDNA) in Saccharomyces cerevisiae cells grown under three different metabolic conditions. The interactions that form between mitochondrial and nuclear loci are dependent on the metabolic state of the yeast. Moreover, the frequency of specific mitochondrial - nuclear interactions (i.e. COX1-MSY1 and Q0182-RSM7) showed significant reductions in the absence of mitochondrial encoded reverse transcriptase machinery. Furthermore, these reductions correlated with increases in the transcript levels of the nuclear loci (MSY1 and RSM7). We propose that these interactions represent an inter-organelle DNA mediated communication system and that reverse transcription of mitochondrial RNA plays a role in this process. Genome Conformation Capture (GCC) has been performed on exponentially growing Saccharomyces cerevisiae cultures in glycerol lactate or galactose media. Paired end sequencing on an Illumina Genome Analyser was performed before the sequences were analysed by the propieatry software Topography 1.19. Inter- and intra- chromosomal interactions were mapped onto the S. cerevisiae S288 genome scaffold.
Project description:Nuclear and mitochondrial organelles must maintain a communication system. Loci on the mitochondrial genome were recently reported to interact with nuclear loci. To determine whether this is part of a DNA based communication system we used genome conformation capture to map the global network of DNA-DNA interactions between the mitochondrial and nuclear genomes (Mito-nDNA) in Saccharomyces cerevisiae cells grown under three different metabolic conditions. The interactions that form between mitochondrial and nuclear loci are dependent on the metabolic state of the yeast. Moreover, the frequency of specific mitochondrial - nuclear interactions (i.e. COX1-MSY1 and Q0182-RSM7) showed significant reductions in the absence of mitochondrial encoded reverse transcriptase machinery. Furthermore, these reductions correlated with increases in the transcript levels of the nuclear loci (MSY1 and RSM7). We propose that these interactions represent an inter-organelle DNA mediated communication system and that reverse transcription of mitochondrial RNA plays a role in this process. Genome Conformation Capture (GCC) has been performed on exponentially growing Saccharomyces cerevisiae cultures in glucose containing media. Paired end sequencing on an Illumina Genome Analyser was performed before the sequences were analysed by the propieatry software Topography 1.19. Inter- and intra- chromosomal interactions were mapped onto the S. cerevisiae S288 genome scaffold.
Project description:Genome-wide copy number changes were monitored using array comparative genomic hybridization (aCGH) of laser-capture microdissected prostate cancer samples spanning stages of prostate cancer progression including precursor lesions, clinically localized disease and metastatic disease. A total of 62 specific cell populations from 38 patients were profiled. Keywords: Disease state analysis using array-based comparatavie genomic hybridization
Project description:Lions and tigers, as dominant apex predators, likely became competitors when lions expanded from Africa into Eurasia approximately one million years ago (Ma), forming a lion–tiger transition belt from the Middle East through Central Asia to the Russian Far East. At the easternmost edge of this zone, the Japanese Archipelago has long been considered a Late Pleistocene tiger refugium, supported by large felid subfossils traditionally attributed to tigers (Panthera tigris), though their taxonomic identity remained unresolved. To clarify the origin, evolutionary history, and biogeography of Japan’s Pleistocene felids, we analyzed 26 ancient specimens previously assumed to be tigers. Using mitochondrial and nuclear genome hybridization capture and sequencing, paleoproteomics, Bayesian molecular dating, and radiocarbon dating, we found that all ancient Japanese “tiger” remains yielding molecular data were, unexpectedly, cave lions (Panthera spelaea). One specimen was radiocarbon dated to 31,060 ± 190 BP. These cave lions likely dispersed to the Japanese Archipelago between ~72.7 and 37.5 thousand years ago (ka), when a land bridge connected northern Japan to the mainland during the Last Glacial Period. Our findings challenge the long-held view that tigers once took refuge in Japan, showing instead that cave lions were widespread in northeast Asia during this period and were the Panthera lineage that colonized Japan, reaching even its southwestern regions despite habitats previously thought to favor tigers.
Project description:Understanding the targeting and spreading patterns of lncRNAs on chromatin requires a technique that can detect both high intensity binding sites and reveal genome-wide spreading patterns with high confidence. We developed an improved hybridization capture protocol to determine lncRNA localization using biotinylated LNA-containing oligonucleotides that hybridize to the target RNA and enhance capture specificity by including a protecting oligonucleotide that competitively displaces contaminating species, leading to highly specific RNA capture. This approach revealed the spreading pattern of roX2, a lncRNA involved in dosage compensation in D. melanogaster, how this pattern relates to chromatin features, and how spreading of roX2 changes upon cellular stress. Upon heat shock, roX2 displays reduced spreading on X chromosome and surprising relocalization to sites on autosomes, revealing how this improved hybridization capture approach can reveal previously uncharacterized changes in the targeting and spreading of lncRNAs on chromatin.
Project description:We provide raw gene sequences of 174 flowering time regulatory genes and gene othologs across a large barley population (895 barley lines) selected from a collection of landrace, cultivated barley, and research varieties of diverse origin. This set represents the whole variety of cultivated barley lifeforms, namely two- and six-row genotypes with winter, spring, and facultative growth habits. We applied a target capture method based on in-solution hybridization using the myBaits® technology (Arbor Biosciences, Ann Arbour, MI, USA) which is based on in-solution biotinylated RNA probes. Baits were designed for flowering time regulatory genes and gene othologs, and used for production of 80mer capture oligonucleotides for hybridization. Genomic DNA was extracted from leaves of a single two-week old barley plant per variety using the cetyl-trimethyl-ammonium bromide (CTAB) method. Physical shearing of genomic DNA was performed with an average size of 275 bp. Library preparation was conducted with KAPA Hyper Prep Kit (KAPA Biosystems, Wilmington, MA). Hybridization of customised RNA baits with capture pools was performed at 65°C for 24 hours. Each pooled sequence capture library was sequenced on an Illumina HiSeq3000 instrument using three lanes to generate paired-end reads per sample. Genome sequencing was conducted at AgriBio, (Centre for AgriBioscience, Bundoora, VIC, Australia).
Project description:Understanding the targeting and spreading patterns of lncRNAs on chromatin requires a technique that can detect both high intensity binding sites and reveal genome-wide spreading patterns with high confidence. We developed an improved hybridization capture protocol to determine lncRNA localization using biotinylated LNA-containing oligonucleotides that hybridize to the target RNA and enhance capture specificity by including a protecting oligonucleotide that competitively displaces contaminating species, leading to highly specific RNA capture. This approach revealed the spreading pattern of roX2, a lncRNA involved in dosage compensation in D. melanogaster, how this pattern relates to chromatin features, and how spreading of roX2 changes upon cellular stress. Upon heat shock, roX2 displays reduced spreading on X chromosome and surprising relocalization to sites on autosomes, revealing how this improved hybridization capture approach can reveal previously uncharacterized changes in the targeting and spreading of a lncRNAs on chromatin.
Project description:Mass spectrometric raw files for "Elucidating the in vivo interactome of HIV-1 RNA by hybridization capture and mass spectrometry."