Return migration has probably been granted the lowest attention in the field of family language policy (FLP). The current paper seeks to address this gap in the research and explores the dynamics of ...FLP of a Polish family in the context of their temporary migration to Germany and return migration to Poland. The authors investigate how mobility affects FLP, especially towards L1 (Polish) and L2 (German) in the context of migration and return migration. The study takes a qualitative, interview-based study design, supported by the language portrait technique. An analysis of the interview data has evidenced the parents’ strong support for the maintenance and development of L1 throughout the whole process of migration and return migration and the lack of it in the case of L2 after the return to Poland. The results have also evidenced that individuals in a family, including children, have significant autonomy and agency and can shape their independ- ent FLPs, which are aligned neither with their parents nor siblings.
Solid frustrated Lewis pair (FLP) catalysts have received much attention. In this work, MOF-808 with rich defects was constructed using a facile monocarboxylic acid modulator-induced defect strategy ...and applied in the transfer hydrogenation of α,β-unsaturated aldehydes to unsaturated alcohol using cyclohexanol as the hydrogen source. MOF-808 was prepared from different zirconium precursors, and it was found that MOF-808-ZT obtained from ZrCl4 and H3BTC forms abundant surface hydroxyl groups (Zr–OH, base sites). Moreover, MOF-808-PA with missing linker defects was constructed by introducing monocarboxylic acid to preoccupy the coordination sites of MOF-808-ZT, leading to abundant Zr–OH2. The coordinated water molecules are removed by the dehydration of MOF-808-PA at 180 °C, and the underlying coordinatively unsaturated Zr4+ (Zr-CUS) is exposed and acts as a Lewis acid site. Characterizations and DFT calculations show that the Zr-CUS/Zr–OH FLPs sites can efficiently activate the CO of aldehydes and −OH of cyclohexanol and reduce their activation energy barrier, thus exhibiting good catalytic performance. Moreover, these FLP sites can easily easily dissociate the H–H bond with a lower activation energy of 0.15 eV, thereby achieving a better catalytic reactivity in the direct hydrogenation of α,β-unsaturated aldehydes to unsaturated alcohols. A possible reaction mechanism was proposed based on in situ DRIFT, vacuum FTIR, and DFT calculations.
Gene stacking by recombinases Srivastava, Vibha; Thomson, James
Plant biotechnology journal,
February 2016, Volume:
14, Issue:
2
Journal Article
Peer reviewed
Open access
Efficient methods of stacking genes into plant genomes are needed to expedite transfer of multigenic traits to crop varieties of diverse ecosystems. Over two decades of research has identified ...several DNA recombinases that carryout efficient cis and trans recombination between the recombination sites artificially introduced into the plant chromosome. The specificity and efficiency of recombinases make them extremely attractive for genome engineering. In plant biotechnology, recombinases have mostly been used for removing selectable marker genes and have rarely been extended to more complex applications. The reversibility of recombination, a property of the tyrosine family of recombinases, does not lend itself to gene stacking approaches that involve rounds of transformation for integrating genes into the engineered sites. However, recent developments in the field of recombinases have overcome these challenges and paved the way for gene stacking. Some of the key advancements include the application of unidirectional recombination systems, modification of recombination sites and transgene site modifications to allow repeated site‐specific integrations into the selected site. Gene stacking is relevant to agriculturally important crops, many of which are difficult to transform; therefore, development of high‐efficiency gene stacking systems will be important for its application on agronomically important crops, and their elite varieties. Recombinases, by virtue of their specificity and efficiency in plant cells, emerge as powerful tools for a variety of applications including gene stacking.
•A FLP/FRT-mediated recombination system is constructed in P. oxalicum.•Double deletion of atf1 and cxrC increases cellulase and xylanase production.•Improved glucose and xylose are released from ...NHSB by engineered Δatf1ΔcxrC::flp.
Penicillium oxalicum has received increasing attention as a potential cellulase-producer. In this study, a copper-controlled flippase recombination enzyme/recognition target (FLP/FRT)-mediated recombination system was constructed in P. oxalicum, to overcome limited availability of antibiotic resistance markers. Using this system, two crucial transcription repressor genes atf1 and cxrC for the production of cellulase and xylanase under solid-state fermentation (SSF) were simultaneously deleted, thereby leading to 2.4- to 29.1-fold higher cellulase and 78.9% to 130.8% higher xylanase production than the parental strain under SSF, respectively. Glucose and xylose released from hydrolysis of pretreated sugarcane bagasse achieved 10.6%–13.5% improvement by using the crude enzymes from the engineered strain Δatf1ΔcxrC::flp under SSF in comparison with that of the parental strain. Consequently, these results provide a feasible strategy for improved cellulase and xylanase production by filamentous fungi.
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•Host-seeking behaviour in Steinernema species correlates with gene expression levels.•The vast majority of microRNAs are unique to each species.•MircoRNAs targetting neuropeptides ...associate with variation in host-seeking behaviour.•Long non-coding isoforms of neuropeptide transcripts may affect microRNA regulation.
We conducted a transcriptomic and small RNA analysis of infective juveniles (IJs) from three behaviourally distinct Steinernema species. Substantial variation was found in the expression of shared gene orthologues, revealing gene expression signatures that correlate with behavioural states. Ninety-seven percent of predicted microRNAs are novel to each species. Surprisingly, our data provide evidence of a new family of non-coding transcripts that overlap with neuropeptide gene loci, which are predicted to influence microRNA regulation of neuropeptide genes. These data suggest that differences in neuropeptide gene expression, isoform variation, and small RNA interactions could contribute to behavioural differences within the Steinernema genus.
Growth and differentiation factor 15 (GDF15) is a stress-induced secreted protein whose plasma levels are increased in obese patients. Recombinant GDF15 reduces body weight (BW) and improves glycemia ...in obese models, which is largely attributed to central action of GDF15 to suppress feeding. Despite this, the tissue specific sites of GDF15 production during obesity are unknown and the potential for local action is understudied. To determine the hepatocyte specific contribution of Gdf15 to changes in circulating GDF15 and effects on obesity, we utilized a Gdf15 KO-first mouse strain where endogenous expression of Gdf15 can be restored in a tissue specific fashion via Flp recombinase (FLP) . Gdf15 KO-first mice received AAV8-TBG-Gfp (KO+GFP) or AAV8-TBG-Flp (liver specific Gdf15 expression; KO+FLP) and were fed HFD for 12 weeks, alongside age-matched C57BL6 mice (WT) . Plasma GDF15 levels were undetectable in KO+GFP and similar between KO+FLP and WT HFD. BW was reduced in KO+FLP compared with KO+GFP and was not different from HFD WT. Interestingly, differences in BW were due to greater changes in lean (LM) compared with fat mass (FM) . Feeding was reduced in KO+FLP when expressed per mouse but not when expressed per g BW or LM, nor when the relationship between BW/LM and feeding were assessed by ANCOVA. Plasma insulin levels were increased in KO+GFP compared with WT HFD and markedly lower in KO+FLP compared with KO+GFP and WT HFD. Hyperinsulinemic euglycemic clamps demonstrated improved insulin sensitivity in KO+FLP due solely to changes in liver insulin action. These data suggest that feeding alone did not account for the changes in BW or insulin resistance, which would favor loss of FM and a generalized improvement in whole-body insulin sensitivity. They also demonstrate that hepatocyte Gdf15 is sufficient for obesity-associated increases in circulating GDF15 and suggest the potential for local effects of GDF15.
Disclosure
B.Xie: None. A.Murali: None. A.Vandevender: None. A.A.Gil silva: None. F.Bello: None. I.J.Sipula: None. N.Dedousis: None. J.Alder: None. M.J.Jurczak: None.
Funding
Pittsburgh Foundation (MR2020 109502)
There are a number of reporter systems that are useful for gene expression analysis in bacteria. However, at least in Salmonella, a versatile and simple luciferase reporter system that can be ...integrated precisely behind a promoter or gene of interest on a chromosome is not currently available. The luciferase operon luxCDABE from Photorhabdus luminescens has several advantages, including brightness, wide linear range, absence in most bacteria, stability at high temperature, and no substrate addition required for the assay. Here, a conjugation-mediated site-specific single-copy luciferase fusion system is developed. A reporter plasmid containing the conditional replication origin R6Kgγ, FRT-luxCDABE, and KmR marker was designed to be incorporated into the FRT site behind the promoter or gene of interest on the chromosome in cells expressing FLP. However, when this reporter plasmid was electroporated directly into such a S. enterica strain, no colonies appeared, likely due to the low transformation efficiency of this relatively large plasmid DNA. Meanwhile, the same reporter plasmid was successfully introduced and launched as an insert of an FRT-containing conjugative transfer plasmid from a mating E. coli strain to the same recipient S. enterica strain, as well as Citrobacter koseri. RcsB-dependent inducible luminescence from the constructed wzc-luxCDABE strains was confirmed. This system is feasible for detecting very low levels of transcription, even in Gram-negative bacterial species that are relatively difficult to genetically manipulate.
Selective hydrogenation of CC double bond in α, β-unsaturated carbonyl compounds is of great significance in the production of fine chemicals, and the design and preparation of hydrogenation ...catalysts with high selectivity is the key to realize the industrialization of this reaction. Controlling and adjusting the steric hindrance between substrate molecules and active center of catalysts is an effective strategy to achieve high selectivity. In this work, the spatially hindered “Frustrated Lewis pairs” (FLPs) was used as the active component and metal-organic frameworks (MOFs) materials with a certain window size were used as the catalyst support. We designed and synthesized a heterogeneous hydrogenation catalyst B(C6F5)3/DO-MIL-101 by immobilizing FLP onto metal-organic frameworks (MIL-101). Characterizations and catalytic performance tests revealed that the catalytically active species B(C6F5)3/DO was confined in MIL-101 nanocavities via Cr–O coordination bond. The resulting B(C6F5)3/DO-MIL-101 catalyst not only realized the recycling of FLP, but also showed excellent catalytic activity and high selectivity in the selective hydrogenation of CC double bond in α, β-unsaturated carbonyl ketones. It should be noted that the steric hindrance between FLP and substrate molecules, as well as the shape selectivity of MOFs nanocages, are the principal factors for the high selectivity of the catalyst.
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•FLP-MOFs heterogeneous catalyst was prepared by post-synthetic modification.•An excellent catalyst for the selective hydrogenation of C＝C double bond in α, β-unsaturated ketones.•The steric effect between FLP and substrates and the shape selectivity of MOFs nanocages were the key to high selectivity.
Several classes of heparan sulfate proteoglycan (HSPG) core proteins and all HS biosynthetic/modifying enzymes are evolutionarily conserved from human to Drosophila melanogaster. This genetically ...tractable model offers highly sophisticated techniques to manipulate gene function in a spatially and temporally controlled manner. Thus, Drosophila genetics has been a powerful system to explore functions of HSPGs in vivo. In this chapter, we will introduce three genetic techniques available in Drosophila: TARGET (temporal and regional gene expression targeting), MARCM (mosaic analysis with a repressible cell marker), and FLP-Out.
ABSTRACT The advent of modern single-cell biology has revealed the striking molecular diversity of cell populations once thought to be more homogeneous. This newly appreciated complexity has made ...intersectional genetic approaches essential to understanding and probing cellular heterogeneity at the functional level. Here, we build on previous knowledge to develop a simple adeno-associated virus (AAV)-based approach to define specific subpopulations of cells by Boolean exclusion logic (AND NOT). This expression by Boolean exclusion (ExBoX) system encodes for a gene of interest that is turned on by a particular recombinase (Cre or FlpO) and turned off by another. ExBoX allows for the specific transcription of a gene of interest in cells expressing only the activating recombinase, but not in cells expressing both. We show the ability of the ExBoX system to tightly regulate expression of fluorescent reporters in vitro and in vivo, and further demonstrate the adaptability of the system by achieving expression of a variety of virally delivered coding sequences in the mouse brain. This simple strategy will expand the molecular toolkit available for cell- and time-specific gene expression in a variety of systems.
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