Serial synchrotron crystallography allows low X‐ray dose, room‐temperature crystal structures of proteins to be determined from a population of microcrystals. Protein production and crystallization ...is a non‐trivial procedure and it is essential to have X‐ray‐compatible sample environments that keep sample consumption low and the crystals in their native environment. This article presents a fast and optimized manufacturing route to metal–polyimide microfluidic flow‐focusing devices which allow for the collection of X‐ray diffraction data in flow. The flow‐focusing conditions allow for sample consumption to be significantly decreased, while also opening up the possibility of more complex experiments such as rapid mixing for time‐resolved serial crystallography. This high‐repetition‐rate experiment allows for full datasets to be obtained quickly (∼1 h) from crystal slurries in liquid flow. The X‐ray compatible microfluidic chips are easily manufacturable, reliable and durable and require sample‐flow rates on the order of only 30 µl h−1.
Serial synchrotron data were collected in flow in a simple microfluidic device under flow‐focusing conditions which allowed sample consumption to be reduced compared with other previously reported liquid‐flow devices. A pulsed data collection strategy prevented the formation of hydrogen bubbles in the chip and the high repetition rate allowed for a short data collection time.
A large proportion of pregnancy losses occur during the pre-implantation period, when the developing embryo is elongating rapidly and signalling its presence to the maternal system. The molecular ...mechanisms that prevent luteolysis and support embryo survival within the maternal environment are not well understood. To gain a more complete picture of these molecular events, genome-wide transcriptional profiles of reproductive day 17 endometrial tissue were determined in pregnant and cyclic Holstein-Friesian dairy cattle.
Microarray analyses revealed 1,839 and 1,189 differentially expressed transcripts between pregnant and cyclic animals (with > or = 1.5 fold change in expression; P-value < 0.05, MTC Benjamini-Hochberg) in caruncular and intercaruncular endometrium respectively. Gene ontology and biological pathway analysis of differentially expressed genes revealed enrichment for genes involved in interferon signalling and modulation of the immune response in pregnant animals.
The maternal immune system actively surveys the uterine environment during early pregnancy. The embryo modulates this response inducing the expression of endometrial molecules that suppress the immune response and promote maternal tolerance to the embryo. During this period of local immune suppression, genes of the innate immune response (in particular, antimicrobial genes) may function to protect the uterus against infection.
Gute Kindesvertretung hat zum Ziel, die Partizipation von Kindern, die in ein juristisches Verfahren involviert sind, zu ermöglichen und mitzugestalten. Die Autor*innen des Bandes stellen diesen ...Prozess erstmals transdisziplinär dar - als Synthese juristischer, psychologischer, (behinderten-)pädagogischer und sozialarbeiterischer Theorie und Praxis. Sie zeichnen die Lage der Kindesvertretung in den deutschsprachigen Ländern nach und entwickeln Standards für alle Phasen der Vertretung, konkretisiert anhand von Fallbeispielen aus der Schweiz. Dabei kommen nicht nur Fachpersonen und Entscheidungsträger*innen zu Wort, sondern auch betroffene Kinder und Jugendliche.
Kindesvertretung Stefan Blum, Sabine Brunner, Peter Grossniklaus, Christophe A. Herzig, Barbara Jeltsch-Schudel, Susanne Meier
2022
eBook
Gute Kindesvertretung hat zum Ziel, die Partizipation von Kindern, die in ein juristisches Verfahren involviert sind, zu ermöglichen und mitzugestalten. Die Autor*innen des Bandes stellen diesen ...Prozess erstmals transdisziplinär dar - als Synthese juristischer, psychologischer, (behinderten-)pädagogischer und sozialarbeiterischer Theorie und Praxis. Sie zeichnen die Lage der Kindesvertretung in den deutschsprachigen Ländern nach und entwickeln Standards für alle Phasen der Vertretung, konkretisiert anhand von Fallbeispielen aus der Schweiz. Dabei kommen nicht nur Fachpersonen und Entscheidungsträger*innen zu Wort, sondern auch betroffene Kinder und Jugendliche.
The functionality of most secreted proteins depends on their assembly into a defined quaternary structure. Despite this, it remains unclear how cells discriminate unassembled proteins en route to the ...native state from misfolded ones that need to be degraded. Here we show how chaperones can regulate and control assembly of heterodimeric proteins, using interleukin 23 (IL-23) as a model. We find that the IL-23 α-subunit remains partially unstructured until assembly with its β-subunit occurs and identify a major site of incomplete folding. Incomplete folding is recognized by different chaperones along the secretory pathway, realizing reliable assembly control by sequential checkpoints. Structural optimization of the chaperone recognition site allows it to bypass quality control checkpoints and provides a secretion-competent IL-23α subunit, which can still form functional heterodimeric IL-23. Thus, locally-restricted incomplete folding within single-domain proteins can be used to regulate and control their assembly.
Female fertility is an important trait in dairy cattle. Identifying putative causal variants associated with fertility may help to improve the accuracy of genomic prediction of fertility. Combining ...expression data (eQTL) of genes, exons, gene splicing and allele specific expression is a promising approach to fine map QTL to get closer to the causal mutations. Another approach is to identify genomic differences between cows selected for high and low fertility and a selection experiment in New Zealand has created exactly this resource. Our objective was to combine multiple types of expression data, fertility traits and allele frequency in high- (POS) and low-fertility (NEG) cows with a genome-wide association study (GWAS) on calving interval in Australian cows to fine-map QTL associated with fertility in both Australia and New Zealand dairy cattle populations. Variants that were significantly associated with calving interval (CI) were strongly enriched for variants associated with gene, exon, gene splicing and allele-specific expression, indicating that there is substantial overlap between QTL associated with CI and eQTL. We identified 671 genes with significant differential expression between POS and NEG cows, with the largest fold change detected for the CCDC196 gene on chromosome 10. Our results provide numerous candidate genes associated with female fertility in dairy cattle, including GYS2 and TIGAR on chromosome 5 and SYT3 and HSD17B14 on chromosome 18. Multiple QTL regions were located in regions with large numbers of copy number variants (CNV). To identify the causal mutations for these variants, long read sequencing may be useful. Variants that were significantly associated with CI were highly enriched for eQTL. We detected 671 genes that were differentially expressed between POS and NEG cows. Several QTL detected for CI overlapped with eQTL, providing candidate genes for fertility in dairy cattle.
BackgroundMany phenotypes in animal breeding are derived from incomplete measures, especially if they are challenging or expensive to measure precisely. Examples include time-dependent traits such as ...reproductive status, or lifespan. Incomplete measures for these traits result in phenotypes that are subject to left-, interval- and right-censoring, where phenotypes are only known to fall below an upper bound, between a lower and upper bound, or above a lower bound respectively. Here we compare three methods for deriving phenotypes from incomplete data using age at first elevation (> 1 ng/mL) in blood plasma progesterone (AGEP4), which generally coincides with onset of puberty, as an example trait.MethodsWe produced AGEP4 phenotypes from three blood samples collected at about 30-day intervals from approximately 5,000 Holstein–Friesian or Holstein–Friesian × Jersey cross-bred dairy heifers managed in 54 seasonal-calving, pasture-based herds in New Zealand. We used these actual data to simulate 7 different visit scenarios, increasing the extent of censoring by disregarding data from one or two of the three visits. Three methods for deriving phenotypes from these data were explored: 1) ordinal categorical variables which were analysed using categorical threshold analysis; 2) continuous variables, with a penalty of 31 d assigned to right-censored phenotypes; and 3) continuous variables, sampled from within a lower and upper bound using a data augmentation approach.ResultsCredibility intervals for heritability estimations overlapped across all methods and visit scenarios, but estimated heritabilities tended to be higher when left censoring was reduced. For sires with at least 5 daughters, the correlations between estimated breeding values (EBVs) from our three-visit scenario and each reduced data scenario varied by method, ranging from 0.65 to 0.95. The estimated breed effects also varied by method, but breed differences were smaller as phenotype censoring increased.ConclusionOur results indicate that using some methods, phenotypes derived from one observation per offspring for a time-dependent trait such as AGEP4 may provide comparable sire rankings to three observations per offspring. This has implications for the design of large-scale phenotyping initiatives where animal breeders aim to estimate variance parameters and estimated breeding values (EBVs) for phenotypes that are challenging to measure or prohibitively expensive.
Quantitative real-time PCR gene expression results are generally normalised using endogenous control genes. These reference genes should be expressed at a constant level across all sample groups in a ...study, and should not be influenced by study treatments or conditions. There has been no systematic investigation of endogenous control genes for bovine endometrium to date. The suitability of both commonly used and novel endogenous control genes was evaluated in this study, with the latter being selected from stably expressed transcripts identified through microarray analysis of bovine endometrium. Fifteen candidate endogenous control genes were assessed across different tissue subtypes in pregnant and cycling Holstein-Friesian dairy cows from two divergent genetic backgrounds.
The expression profiles of five commonly used endogenous control genes (GAPDH, PPIA, RPS9, RPS15A, and UXT) and 10 experimentally derived candidate endogenous control genes (SUZ12, C2ORF29, ZNF131, ACTR1A, HDAC1, SLC30A6, CNOT7, DNAJC17, BBS2, and RANBP10) were analysed across 44 samples to determine the most stably expressed gene. Gene stability was assessed using the statistical algorithms GeNorm and Normfinder. All genes presented with low overall variability (0.87 to 1.48% CV of Cq). However, when used to normalise a differentially expressed gene (oxytocin receptor - OXTR) in the samples, the reported relative gene expression levels were significantly affected by the control gene chosen. Based on the results of this analysis, SUZ12 is proposed as the most appropriate control gene for use in bovine endometrium during early pregnancy or the oestrus cycle.
This study establishes the suitability of novel endogenous control genes for comparing expression levels in endometrial tissues of pregnant and cycling bovines, and demonstrates the utility of microarray analysis as a method for identifying endogenous control gene candidates.
The development of biomarkers of fertility could provide benefits for the genetic improvement of dairy cows. Circulating small extracellular vesicles (sEVs) show promise as diagnostic or prognostic ...markers since their cargo reflects the metabolic state of the cell of origin; thus, they mirror the physiological status of the host. Here, we employed data-independent acquisition mass spectrometry to survey the plasma and plasma sEV proteomes of two different cohorts of Young (Peripubertal; n = 30) and Aged (Primiparous; n = 20) dairy cows (Bos taurus) of high- and low-genetic merit of fertility and known pregnancy outcomes (ProteomeXchange data set identifier PXD042891). We established predictive models of fertility status with an area under the curve of 0.97 (sEV; p value = 3.302e-07) and 0.95 (plasma; p value = 6.405e-08). Biomarker candidates unique to high-fertility Young cattle had a sensitivity of 0.77 and specificity of 0.67 (*p = 0.0287). Low-fertility biomarker candidates uniquely identified in sEVs from Young and Aged cattle had a sensitivity and specificity of 0.69 and 1.0, respectively (***p = 0.0005). Our bioinformatics pipeline enabled quantification of plasma and circulating sEV proteins associated with fertility phenotype. Further investigations are warranted to validate this research in a larger population, which may lead to improved classification of fertility status in cattle.
The 2-year lidar water vapor database (November 2013 - October 2015) of the Maïdo Observatory (Reunion Island / 21°S,55.5°E) is now processed. The performances of the lidar in providing accurate ...vertical structures are shown to be good. The ability to measure quantities of a few ppmv in the lower stratosphere is demonstrated (based on Cryogenic Frost point Hygrometer sonde/lidar profiles comparisons) for a 48-hour integration time period, up to 22 km (with a vertical resolution of 1.3 km).
PDF
