Defective phage formation by lysogens of integration deficient phage P22 mutants

Integration deficient (L) mutants of phage P22 can be complemented with L+ phage to yield L mutant lysogens. Once established the L prophage is stable in the absence of L+ function. Ultraviolet or thermal induction of L lysogens leads to production of defective phage particles and low yields of infectious phage. The defective particles contain a normal amount of DNA but the DNA is mainly bacterial in origin and appears to derive from the pro C region adjacent to one end of the prophage. The defective lysates transduce pro C at high frequency relative to other bacterial markers. The representation of phage genes is strongly polar with markers distal to pro C being rarely present. It appears that L+ function is required not only for efficient integration of the prophage, but for normal recovery of the prophage genome following induction.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/33221/1/0000611.pd

A phage P22 gene controlling integration of prophage

A mutually noncomplementing group of phage P22 mutants (L mutants) have been isolated which cannot accomplish a late step necessary for prophage integration. L mutants produce turbid plaques and lead to cell survival on high multiplicity infection, but the infecting phage genomes are incapable of integrating as prophage and are progressively diluted among the segregating cells. L mutants can become prophage by complementation with L+ phage. The L+ gene apparently produces a diffusible product which acts trans to accomplish integration. Experiments with temperature sensitive L mutants show that the gene begins acting late, after the infected cells have recovered and begun to divide, and does not accomplish its effect in all the cells until at least the third generation. L lysogens formed by complementation are stable. The L+ gene is thus necessary for integration but not for maintenance of the lysogenic state.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/33387/1/0000786.pd

The synthesis of phage and host dna in the establishment of lysogeny

A method of extraction is described which prevents excessive fragmentation of bacterial DNA. The larger bacterial DNA can then be separated from the phage DNA on sucrose gradients. The relative amounts of phage and host DNA made at various times during infection of Salmonella typhimurium with phage P22 leading to lysogeny have been determined. Phage-specific DNA synthesis begins at about 4 minutes, continues to increase for 2-4 minutes, and then decreases until complete repression is attained at 16 minutes. Autonomous phage DNA synthesis remains repressed from then on. Bacterial-specific synthesis proceeds at a decreasing rate until about 16 minutes, when almost complete inhibition is reached. When DNA synthesis recovers in the infected cells, only host-specific material is made. A new species of DNA, which is made only when phage DNA is synthesized, has been detected. The possible implications of these findings in the understanding of the lysogeny interaction is discussed.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32039/1/0000082.pd

Overexpression of an endogenous type 2 diacylglycerol acyltransferase in the marine diatom Phaeodactylum tricornutum enhances lipid production and omega-3 long-chain polyunsaturated fatty acid content

Background: Oleaginous microalgae represent a valuable resource for the production of high-value molecules. Considering the importance of omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs) for human health and nutrition the yields of high-value eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) require significant improvement to meet demand; however, the current cost of production remains high. A promising approach is to metabolically engineer strains with enhanced levels of triacylglycerols (TAGs) enriched in EPA and DHA. Results: Recently, we have engineered the marine diatom Phaeodactylum tricornutum to accumulate enhanced levels of DHA in TAG. To further improve the incorporation of omega-3 LC-PUFAs in TAG, we focused our effort on the identification of a type 2 acyl-CoA:diacylglycerol acyltransferase (DGAT) capable of improving lipid production and the incorporation of DHA in TAG. DGAT is a key enzyme in lipid synthesis. Following a diatom based in vivo screen of candidate DGATs, a native P. tricornutum DGAT2B was taken forward for detailed characterisation. Overexpression of the endogenous P. tricornutum DGAT2B was confirmed by qRT-PCR and the transgenic strain grew successfully in comparison to wildtype. PtDGAT2B has broad substrate specificity with preferences for C16 and LC-PUFAs acyl groups. Moreover, the overexpression of an endogenous DGAT2B resulted in higher lipid yields and enhanced levels of DHA in TAG. Furthermore, a combined overexpression of the endogenous DGAT2B and ectopic expression of a Δ5-elongase showed how iterative metabolic engineering can be used to increase DHA and TAG content, irrespective of nitrogen treatment. Conclusion: This study provides further insight into lipid metabolism in P. tricornutum and suggests a metabolic engineering approach for the efficient production of EPA and DHA in microalgae

Designer diatom episomes delivered by bacterial conjugation.

Eukaryotic microalgae hold great promise for the bioproduction of fuels and higher value chemicals. However, compared with model genetic organisms such as Escherichia coli and Saccharomyces cerevisiae, characterization of the complex biology and biochemistry of algae and strain improvement has been hampered by the inefficient genetic tools. To date, many algal species are transformable only via particle bombardment, and the introduced DNA is integrated randomly into the nuclear genome. Here we describe the first nuclear episomal vector for diatoms and a plasmid delivery method via conjugation from Escherichia coli to the diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana. We identify a yeast-derived sequence that enables stable episome replication in these diatoms even in the absence of antibiotic selection and show that episomes are maintained as closed circles at copy number equivalent to native chromosomes. This highly efficient genetic system facilitates high-throughput functional characterization of algal genes and accelerates molecular phytoplankton research

Associations of cognitive reserve and psychological resilience with cognitive functioning in subjects with cerebral white matter hyperintensities

Background and purpose Cerebral small vessel disease is characterized by progressive white matter hyperintensities (WMH) and cognitive decline. However, variability exists in how individuals maintain cognitive capabilities despite significant neuropathology. The relationships between individual cognitive reserve, psychological resilience and cognitive functioning were examined in subjects with varying degrees of WMH. Methods In the Helsinki Small Vessel Disease Study, 152 subjects (aged 65-75 years) underwent a comprehensive neuropsychological assessment, evaluation of subjective cognitive complaints and brain magnetic resonance imaging with volumetric WMH evaluation. Cognitive reserve was determined by education (years) and the modified Cognitive Reserve Scale (mCRS). Psychological resilience was evaluated with the Resilience Scale 14. Results The mCRS total score correlated significantly with years of education (r = 0.23, p < 0.01), but it was not related to age, sex or WMH volume. Together, mCRS score and education were associated with performance in a wide range of cognitive domains including processing speed, executive functions, working memory, verbal memory, visuospatial perception and verbal reasoning. Independently of education, the mCRS score had incremental predictive value on delayed verbal recall and subjective cognitive complaints. Psychological resilience was not significantly related to age, education, sex, WMH severity or cognitive test scores, but it was associated with subjective cognitive complaints. Conclusions Cognitive reserve has strong and consistent associations with cognitive functioning in subjects with WMH. Education is widely associated with objective cognitive functioning, whereas lifetime engagement in cognitively stimulating leisure activities (mCRS) has independent predictive value on memory performance and subjective cognitive complaints. Psychological resilience is strongly associated with subjective, but not objective, cognitive functioning.Peer reviewe

Proceedings of the 2015 WA Chapter of MSA Symposium on Music Performance and Analysis

This publication, entitled Proceedings of the 2015 WA Chapter MSA Symposium on Music Performance and Analysis, is a double-blind peer-reviewed conference proceedings published by the Western Australian Chapter of the Musicological Society of Australia, in conjunction with the Western Australian Academy of Performing Arts, Edith Cowan University, edited by Jonathan Paget, Victoria Rogers, and Nicholas Bannan. The original symposium was held at the University of Western Australia, School of Music, on 12 December 2015. With the advent of performer-scholars within Australian Universities, the intersections between analytical knowledge and performance are constantly being re-evaluated and reinvented. This collection of papers presents several strands of analytical discourse, including: (1) the analysis of music recordings, particularly in terms of historical performance practices; (2) reinventions of the \u27page-to-stage\u27 paradigm, employing new analytical methods; (3) analytical knowledge applied to pedagogy, particularly concerning improvisation; and (4) so-called \u27practice-led\u27 research.https://ro.ecu.edu.au/ecubooks/1005/thumbnail.jp