Biology subjects

Pampuch, M.

Publications and source records attributed to Pampuch, M..

3 recordsLinked to original sources

bioRxiv2026

Cultivation and physiological characterization of a desert-derived Halospirulina isolate

Here, we describe a filamentous Halospirulina isolate (Halospirulina saudiensis) obtained from water-clay microhabitat the Empty Quarter desert, (ar-Rub al-Kh[a]l[i]), Saudi Arabia which grows in saline conditions. We present its fully sequenced genome, the first for the genus, and characterize its growth dynamics as well as biochemical composition under a range of cultivation conditions. Protein, carbohydrate, lipid, and phycocyanin content varied with cultivation regime but were largely stable. H. saudiensis reached biomass concentrations of up to 9.83 g L-1 at pH 7, 35 {degrees}C and continuous 325 {micro}mol photons m-2 s-1. Variable climate simulations in lab-scale photobioreactors revealed preference for warmer season cultivation under modeled outdoor conditions. Carotenoid analysis revealed a pigment profile enriched in canthaxanthin and other ketocarotenoids, distinguishing it from industrial Limnospira and positioning its value for neutraceuticals and feed additives. Genome analysis identified a carotene ketolase (crtO) homolog consistent with other cyanobacteria that accumulate ketocarotenoids. Phycocyanin content was heavily dependent on culture health and varied with cultivation pH, irradiance, reaching maximum values of 67.3 {+/-} 0.8 mg gDW-1 (6.73 %). Extracted phycocyanin showed marginal thermal stability compared to that from L. platensis. The findings suggest that H. saudiensis could be a promising source of biomass, ketocarotenoids, and natural pigments, cultivated in saline conditions with elevated temperature and irradiance. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=71 SRC="FIGDIR/small/728284v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@17d6143org.highwire.dtl.DTLVardef@7cb1f7org.highwire.dtl.DTLVardef@880df4org.highwire.dtl.DTLVardef@4c16a6_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIHalospirulina saudiensis sp. nov. isolated from Empty Quarter C_LIO_LIFirst genome-resolved characterization of a Halospirulina strain C_LIO_LIReached 9.83 g L-1 in Red Sea salinity conditions C_LIO_LIAccumulates canthaxanthin as major carotenoid C_LIO_LIPhycocyanin slightly thermotolerant C_LI

microbiology↗

bioRxiv2024

Spheroplasted cells: a game changer for DNA delivery to diatoms

Diatoms produce 20% of the worlds fixed organic carbon annually, making them vital to global carbon fixation and climate change mitigation. Their potential as cell factories for biofuels, proteins, and other high value chemicals remains underutilized due to a lack of genetic engineering tools, with DNA delivery being one of the biggest challenges. Here, we present optimized electroporation and polyethylene glycol transformation methods for delivering DNA and ribonucleoprotein complexes to Phaeodactylum tricornutum, a model diatom species and emerging chassis for algal biotechnology. It was possible to recover transformants with as little as 1 ng of DNA, and to transform linear or circular episomes as large as 55.6 kb. With the optimized electroporation protocol, episomes can be assembled in the algal cell de novo through diatom in vivo assembly (DIVA), forgoing the need for time-consuming traditional cloning steps in Escherichia coli and Saccharomyces cerevisiae. It was also possible to electroporate a Cas9 ribonucleoprotein complex in P. tricornutum, providing an alternative to biolistics for DNA free genome engineering. We have demonstrated that the PEG approach can be adapted to successfully transform Thalassiosira pseudonana, demonstrating the applicability of our methods for engineering other diatom species. These tools can be used to accelerate diatom synthetic biology projects and, therefore, the development of sustainable technologies.

synthetic biology↗

bioRxiv2023

Design and assembly of the 117-kb Phaeodactylum tricornutum chloroplast genome

There is a growing impetus to expand the repository of chassis available to synthetic biologists. The chloroplast genome presents a unique chassis for engineering photosynthetic eukaryotes by virtue of its compact size, lack of epigenetic regulation, and containment within the secluded lipid bilayers of the organelle. The development of the chloroplast as a synthetic biology chassis, however, has been limited by a lack of efficient techniques for whole genome cloning and engineering. Here, we demonstrate two approaches for cloning the 117 kb Phaeodactylum tricornutum chloroplast genome that have 90 to 100% efficiency when screening as few as ten Saccharomyces cerevisiae colonies following yeast assembly. The first method directly uses PCR-amplified fragments of the genome for yeast assembly, whereas the second method relies upon the pre-cloning of eight overlapping genomic regions into individual plasmids that they can later be released from. The cloned genome can be stably maintained and propagated within Escherichia coli, which provides an exciting opportunity for engineering a novel delivery mechanism for bringing DNA directly to the algal chloroplast. As well, one of the cloned genomes was designed to contain a single SapI site within the yeast URA3 open-reading frame, which can be used to linearize the genome and integrate designer cassettes via golden-gate cloning or further iterations of yeast assembly.

synthetic biology↗