Biology subjects

Bischof, K.

Publications and source records attributed to Bischof, K..

3 recordsLinked to original sources

bioRxiv2026

A synthetic biology approach to bacterial transcription initiation: RNA aptamer based in vitro transcription assay for rapidly testing bacterial RNA polymerases, promoters and inhibitors.

We present a robust and versatile in vitro transcription (IVT) assay based on an optimized Broccoli RNA aptamer sequence. When paired with the fluorophore DFHBI-1T, this system enables real-time monitoring of multi-round transcription over several hours. To facilitate streamlined promoter analysis, we developed the pIVT3 plasmid backbone. The system was validated using both the single-subunit T7 RNA polymerase and the multi-subunit Escherichia coli RNA polymerase; notably, the activity of the E. coli enzyme remained strictly dependent on the presence of a {sigma} factor and a cognate promoter. To optimize the signal-to-noise ratio, we incorporated two rrnBT1 terminators upstream of the promoter of interest. This modification effectively eliminated background transcription for weak promoters (PlivJ) and prevented interference from read-through transcription in strong synthetic promoters (Ptrc*). Furthermore, we demonstrated the assays utility for drug discovery by characterizing the time- and dose-dependent inhibitory kinetics of rifampicin. Collectively, these results establish the Broccoli-based IVT system as a highly adaptable platform for quantifying promoter strength and screening small-molecule inhibitors of bacterial transcription. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/744185v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1632360org.highwire.dtl.DTLVardef@1a77977org.highwire.dtl.DTLVardef@1780c9org.highwire.dtl.DTLVardef@1082e05_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2022

Chloroplast DNA methylation in the kelp Saccharina latissima is determined by origin and influenced by cultivation

DNA cytosine methylation is an important epigenetic mechanism in genomic DNA, but absent in the chloroplast DNA of most land plants. We detected methylation in the chloroplast DNA of the kelp Saccharina latissima, a non-model macroalgal species of high ecological (wild populations) and economic importance. Since the functional role of the chloroplast methylome is yet largely unknown, we compared for the first time the chloroplast DNA cytosine methylation between wild and cultured kelp from different climatic origins (High- Arctic (79 {degrees}N) and temperate (54 {degrees}N), laboratory samples at 5 {degrees}C, 10 {degrees}C and 15 {degrees}C). Our results suggest genome-wide differences in methylated sites, and methylation level, between the origins, and a strong effect of cultivation. Functions related to photosynthesis showed differential methylation only between origins. Significant differences in methylation between cultivated and wild samples of genes related to transcription and translation were unique to the high-arctic samples. Both findings indicate that origin and cultivation strongly, but differently affect the chloroplast methylome. Similar methylomes for samples from the same origin - independent from whether they grow in the wild or in the lab - suggest that origin- specific methylation marks on the chloroplast genome are inherited. Given that DNA- methylation affects gene expression, our study suggests that lab-cultivation alters epigenetically determined kelp chloroplast characteristics at least to the same degree as ecotypic differentiation does, but likely on a much shorter timescale. This indicates the capacity for rapid non-genetic adaptation in the kelp Saccharina latissima.

genomics↗

bioRxiv2019

Temperature dependent sex-biased gene expression in the gametophytes of the kelp Saccharina latissima

Saccharina latissima is an economically and ecologically relevant kelp species in Europe and North America. In kelps, the sexuality is expressed during the haploid life stage and the microscopic gametophytes exhibit significant sexual dimorphism. To understand the sex-dependent impact of temperature on the gametophyte stage, we analyzed for the first time, gene expression profiles of male and female gametophytes at three different temperatures (4{degrees}C, 12{degrees}C and 20{degrees}C) characteristic for the species distribution range by using RNA-sequencing. We identified several differentially expressed genes between sexes; while female biased genes were enriched in general metabolism and energy production, male biased genes function within cell cycle and signaling. In our study, temperature modulated sex-biased gene expression, with only a small percentage of differentially expressed genes consistently male (7%) or female-biased (12%) at the three temperatures. Female gametophytes responded stronger to higher temperatures than males, suggesting that males are more heat tolerant. Differences between S. latissima and other brown algal gender-dependent gene expression might mirror the different evolutionary and ecological contexts. Genomic information on kelp gametophyte is still scarce and thus this study adds to our knowledge on sex differences in abiotic stress responses in macroalgae at the transcriptomic level.\n\nHighlightThe transcriptomic basis for sexual dimorphism and associated metabolic needs are described for the kelp Saccharina latissima. Temperature modulates sex-biased gene expression resulting in a stronger stress response in females.

plant biology↗