Biology subjects

Trau, M.

Publications and source records attributed to Trau, M..

6 recordsLinked to original sources

bioRxiv2025

Tracking breast cancer progression using Methylscape

Cancer progression is intricately driven by epigenetic reprogramming, where aberrant DNA methylation patterns reshape gene regulation and cellular phenotypes. These alterations include promoter hypermethylation of tumour suppressor genes and global hypomethylation, collectively fuelling oncogenesis and disease advancement. We previously introduced the Methylscape--a distinct cancer-specific DNA methylation landscape characterized by clustered promoter hypermethylation and gene body hypomethylation--that enhances DNAs physical affinity for gold surfaces. In this study, we demonstrate that Methylscape can be leveraged to monitor cancer progression. Using a breast cancer epithelial-mesenchymal transition (EMT) model, we observe an increase in Methylscape enrichment of mesenchymal-state DNA during EMT, suggesting that this method can sensitively detect subtle epigenetic remodelling linked to tumour progression. Using a gold-based DNA desorption enrichment strategy coupled with methylation sequencing and qPCR, we show that hypermethylated regions are preferentially enriched. Finally, we developed a low-cost, disposable screen-printed electrode platform for stage-specific breast cancer monitoring. Together, these findings establish Methylscape as a promising biophysical biomarker for non-invasive and real-time monitoring of cancer progression, advancing its potential for clinical translation.

bioengineering↗

bioRxiv2024

DIYNAFLUOR: An Affordable DIY Plug-and-Play Nucleic Acid Fluorometer for eDNA Quantification in Resource Limited Settings

Nucleic Acid (NA) fluorometry is widely employed for quantifying environmental DNA (eDNA) samples and their downstream DNA sequencing libraries, owing to its sensitivity, accuracy, and speed. However, the high cost of NA fluorometers presents a barrier to eDNA sequencing in resource limited settings (RLSs). For instance, at [~]$1.5-3.3k USD, current NA fluorometers present a greater capital cost than ONTs $1k USD portable MinION third-generation Nanopore sequencing platform. The collapse of international scientific device and consumable supply chains during the COVID-19 pandemic also highlighted the need for distributed manufacturing of molecular research tools to mitigate the impact of increased pricing to RLSs. To address these challenges, we have developed the "DIYNAFLUOR" (DIY Nucleic Acid FLUORometer), a portable, open-source, <$40 USD NA fluorometer, designed using readily available off-the-shelf components, simple 3D-printed parts, and plug-and-play, solder-free assembly. The DIYNAFLUOR was primarily designed to be compatible with the popular DNA-centric Qubit High Sensitivity (HS) and Broad Range (BR) assay kits. Notably, the DIYNAFLUOR demonstrated an in-assay Limit of Detection with the Qubit HS kit of 0.0028 ng/L, and an average absolute bias of 0.018 ng/L across a 0-10 ng/L working range using a 2-point linear calibration methodology. Device verification was performed by comparative measurements with a Qubit 4 fluorometer in a busy biotechnology laboratory, and build instructions were validated through assembly and qualification of three DIYNAFLUOR devices by researchers outside the primary design team. We also describe a custom "extreme"low-cost assay, <13{cents} USD per-measurement, that uses SYBR Safe dye to quantify DNA across a working range of 0-0.5 ng/L. This assay reports a lower sensitivity and accuracy than commercial kits but may be of use to RLSs in times of extreme resource constraints or as a teaching tool for STEM educators. To demonstrate its practical application for field-based eDNA analysis in RLSs, the DIYNAFLUOR was used to perform all quality control measurements throughout the preparation of a 16S and 18S metabarcoding library generated from eDNA extracted from Australian lake water, leading to the successful identification of Australian fauna via Nanopore sequencing. Finally, configurations of the DIYNAFLUOR for RNA and Protein quantification are briefly described. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=70 SRC="FIGDIR/small/626200v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@28244dorg.highwire.dtl.DTLVardef@16749acorg.highwire.dtl.DTLVardef@15c31aorg.highwire.dtl.DTLVardef@bc03e8_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Unraveling cfDNA Fragmentation Patterns: Linking Chromatin Architecture to Cancer Diagnostic Sensitivity

Circulating cell-free DNA (cfDNA) liquid biopsies offer a powerful, non-invasive approach for cancer diagnostics, leveraging DNA fragments primarily released during apoptosis. Using droplet digital PCR on cfDNA samples from 88 breast and colorectal cancer patients, we uncovered highly consistent nucleosome breakpoints across individuals. Proximity to these cfDNA breakpoints diminishes the sensitivity of somatic mutation assays, including those targeting IDH1 and NRAS hotspot mutations, underscoring the importance of precise nucleosome positioning data. We introduce a probabilistic nucleosome protection scoring method that enables high-resolution mapping of nucleosome positions and cfDNA breakpoints. These maps reveal distinct nucleosome spacing patterns between heterochromatin and euchromatin, corresponding to different chromatin fiber topologies. Analysis of X-chromosome inactivation further supports the topoisomer model: heterochromatins 187 bp nucleosome repeat length differs from euchromatins 182 bp, generating wave interference patterns in nucleosome signals. Our findings suggest that loop-mediated transitions from T2 to T1 topoisomers shape chromatin accessibility and cfDNA fragmentation, advancing the diagnostic potential of liquid biopsy assays. HighlightsO_LINucleosome breakpoint conservation impacts somatic mutation assay sensitivity C_LIO_LIProbabilistic protection scoring enhances cfDNA nucleosome peak resolution C_LIO_LIChromatin states exhibit unique nucleosome spacing linked to distinct topologies, influencing cfDNA fragmentation C_LIO_LIChromosome X-inactivation phasing analyses reveal waveform interference in nucleosome positioning signals C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/623159v1_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@65893corg.highwire.dtl.DTLVardef@14c986org.highwire.dtl.DTLVardef@ba9306org.highwire.dtl.DTLVardef@10a7364_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2024

Cardiovascular symptoms of PASC are associated with trace-level cytokines that affect the function of human pluripotent stem cell derived cardiomyocytes

Globally, over 65 million individuals are estimated to suffer from post-acute sequelae of COVID-19 (PASC). A large number of individuals living with PASC experience cardiovascular symptoms (i.e. chest pain and heart palpitations) (PASC-CVS). The role of chronic inflammation in these symptoms, in particular in individuals with symptoms persisting for >1 year after SARS-CoV-2 infection, remains to be clearly defined. In this cross-sectional study, blood samples were obtained from three different sites in Australia from individuals with i) a resolved SARS-CoV-2 infection (and no persistent symptoms i.e. Recovered), ii) individuals with prolonged PASC-CVS and iii) SARS-CoV-2 negative individuals. Individuals with PASC-CVS, relative to Recovered individuals, had a blood transcriptomic signature associated with inflammation. This was accompanied by elevated levels of pro-inflammatory cytokines (IL-12, IL-1{beta}, MCP-1 and IL-6) at approximately 18 months post-infection. These cytokines were present in trace amounts, such that they could only be detected with the use of novel nanotechnology. Importantly, these trace-level cytokines had a direct effect on the functionality of pluripotent stem cell derived cardiomyocytes in vitro. This effect was not observed in the presence of dexamethasone. Plasma proteomics demonstrated further differences between PASC-CVS and Recovered patients at approximately 18 months post-infection including enrichment of complement and coagulation associated proteins in those with prolonged cardiovascular symptoms. Together, these data provide a new insight into the role of chronic inflammation in PASC-CVS and present nanotechnology as a possible novel diagnostic approach for the condition.

microbiology↗

bioRxiv2024

The integration of tandem gene repeats via a bacterial type-II toxin-antitoxin-mediated gene amplification (ToxAmp) system and stability visualisation in Saccharomyces cerevisiae

Tandem gene repeats naturally occur as important genomic features and determine many traits in living organisms, like human diseases and microbial productivities of target bioproducts. Here, we develop a bacterial type-II toxin-antitoxin-mediated method to manipulate genomic integration of tandem gene repeats in Saccharomyces cerevisiae and further visualise the evolutionary trajectories of gene repeats. We designed a tri-vector system to introduce toxin-antitoxin-driven gene amplification (ToxAmp) modules, and accidentally re-visited the high-level capacity of multi-fragment co-transformation in S. cerevisiae. This system delivered the multi-copy gene integration in the form of tandem gene repeats spontaneously and independently from toxin-antitoxin-mediated selection. Inducing the toxin (RelE) expressing via a copper (II)-inducible CUP1 promoter successfully drove the in-situ gene amplification of the antitoxin (RelB) module, resulting in [~]40 copies of a green fluorescence reporter (GFP) gene per copy of genome. The copy-number changes, increasing and decreasing, and stable maintenance were visualised using the GFP and blue chromoprotein AeBlue as reporters. Copy-number increasing happened spontaneously not depending on a selection pressure and was quickly enriched through toxin-antitoxin-mediated selection. In summary, the bacterial toxin-antitoxin systems provide a flexible mechanism to manipulate gene copy number in eukaryotic cells and can be exploited for synthetic biology and metabolic engineering applications. Table of Contents Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/578080v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@75d1f3org.highwire.dtl.DTLVardef@12aafeforg.highwire.dtl.DTLVardef@184a1a5org.highwire.dtl.DTLVardef@90586a_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2021

An analytical pipeline for DNA Methylation Array Biomarker Studies

DNA methylation is one of the most commonly studied epigenetic biomarkers, due to its role in disease and development. The Illumina Infinium methylation arrays still remains the most common method to interrogate methylation across the human genome, due to its capabilities of screening over 480, 000 loci simultaneously. As such, initiatives such as The Cancer Genome Atlas (TCGA) have utilized this technology to examine the methylation profile of over 20,000 cancer samples. There is a growing body of methods for pre-processing, normalisation and analysis of array-based DNA methylation data. However, the shape and sampling distribution of probe-wise methylation that could influence the way data should be examined was rarely discussed. Therefore, this article introduces a pipeline that predicts the shape and distribution of normalised methylation patterns prior to selection of the most optimal inferential statistics screen for differential methylation. Additionally, we put forward an alternative pipeline, which employed feature selection, and demonstrate its ability to select for biomarkers with outstanding differences in methylation, which does not require the predetermination of the shape or distribution of the data of interest. AvailabilityThe Distribution test and the feature selection pipelines are available for download at: https://github.com/uqjlu8/DistributionTest

bioinformatics↗