Probabilistic base calling of Solexa sequencing data

Background  

Solexa/Illumina short-read ultra-high throughput DNA sequencing technology produces millions of short tags (up to 36 bases) by parallel sequencing-by-synthesis of DNA colonies. The processing and statistical analysis of such high-throughput data poses new challenges; currently a fair proportion of the tags are routinely discarded due to an inability to match them to a reference sequence, thereby reducing the effective throughput of the technology.     

Choice of scan-sampling intervals—An example with quantifying neighbours in dairy cows

To date, only a few studies using scan-sampling intervals to record neighbours in groups of animals explain how the interval length was chosen. In this study, we investigated the effects of different scan-sampling intervals, behavioural context, and the definition of neighbours on the accuracy of data as compared to a quasi-continuous observation. The data was collected from dairy cows kept in cubicle housing systems. Dairy herds of 22–43 cows on 6 farms were observed once a minute with an automatic tracking system for 6 days, and both the nearest neighbour and all neighbours within a defined distance of each cow were recorded with references to the activity, feeding and lying area and stored in neighbourship matrices. In the analysis, we used data collected at intervals of 2, 3, 4, …, 30 min, and correlated this simulated data with the data based on the 1-min interval using a specialised correlation coefficient for matrices τ_Kr. This correlation coefficient was then used as the response variable in mixed-effects models. We found that the size of the correlation coefficients generally decreased as interval length increased. This decrease was less pronounced for all neighbours than for the nearest neighbour. Moreover, the decrease was greatest with data from the activity area and lowest with data from the lying area. We concluded that, even with the relatively slow dairy cows in a barn environment, neighbour recordings should be conducted at short scan-sampling intervals in order to achieve a minimum correlation of τ_Kr = 0.8 with the quasi-continuous data. Intervals of every 2, 8 and 17 min are recommended for observation of neighbours in dairy cows for the activity, feeding and lying areas, respectively, and species that move faster may well require even shorter sampling intervals.     

Differences between single and paired heifers in residency in functional areas, length of travel path, and area used throughout days 1–6 after integration into a free stall dairy herd

The integration of heifers into free stall dairy herds is a frequent management procedure, but little systematic research has been conducted on its effect on cow behavior. Previous studies mainly focused on aggressive interactions, but it is also of interest how integration affects the spatial distribution of both the cows in the herd and the integrated heifers. In the present study we integrated a single and a pair of heifers on each of six Swiss working farms in a balanced order. Using an automatic tracking system, we recorded the positions of all the cows and of the integrated heifers at 1 min intervals for six continuous 24 h periods. From these data we calculated the proportion of time the animals spent in the activity area, at the feed rack and in the lying cubicles, their average path length and the area of the barn that they used. We then compared the behavior of the integrated heifers with that of the cows in the introductory weeks. We also compared the behavior of the cows recorded in the control weeks directly preceding the integration and in the introductory weeks. For evaluation we used linear mixed-effects models. Singly integrated heifers spent a higher proportion of time in the activity area (0.29 vs. 0.14; P < 0.001) and a lower proportion of time in the lying area (0.40 vs. 0.53; interaction with day, P = 0.011) than the cows, whereas the heifers of the pairs mainly spent a lower proportion of time in the feeding area than the cows (0.23 vs. 0.32; interaction with day, P = 0.044). Average path length was longer for the integrated heifers soon after introduction but approached the values of the cows later on (interaction with day, P = 0.012). The total barn area used by a given animal was largest in the cows and was reduced in heifers integrated singly or in pairs (cows: 341/373 m², pairs: 306 m², single heifer: 333 m²; P = 0.055). Cows were little influenced in their space use by the integration of a single or pair of heifers. In summary, the behavior of the singly integrated heifers differed more markedly from that of the cows than the behavior of the heifers introduced in pairs during the introductory week. We would therefore recommend integrating pairs rather than single heifers into herds of dehorned dairy cows to ease their integration.     

Independent evolution of functional Pm3 resistance genes in wild tetraploid wheat and domesticated bread wheat

The Pm3 alleles of cultivated bread wheat confer gene for gene resistance to the powdery mildew fungus. They represent a particular case of plant disease resistance gene evolution, because of their recent origin and possible evolution after the formation of hexaploid wheat. The Pm3 locus is conserved in tetraploid wheat, thereby allowing the comparative evolutionary study of the same resistance locus in a domesticated species and in one of its wild ancestors. We have identified 61 Pm3 allelic sequences from wild and domesticated tetraploid wheat subspecies. The Pm3 sequences corresponded to 24 different haplotypes. They showed low sequence diversity, differing by only a few polymorphic sequence blocks that were further reshuffled between alleles by gene conversion and recombination. Polymorphic sequence blocks are different from the blocks found in functional Pm3 alleles of hexaploid wheat, indicating an independent evolution of the Pm3 loci in the two species. A new functional gene was identified in a wild wheat accession from Syria. This gene, Pm3k , conferred intermediate race-specific resistance to powdery mildew, and consists of a mosaic of gene segments derived from non-functional alleles. This demonstrates that Pm3 -based resistance is not very frequent in wild tetraploid wheat, and that the evolution of functional resistance genes occurred independently in wild tetraploid and bread wheat. The Pm3 sequence variability and geographic distribution indicated that diversity was higher in wild emmer wheat from the Levant area, compared with the accessions from Turkey. Further screens for Pm3 functional genes in wild wheat should therefore focus on accessions from the Levant region.     

New classes of potent and bioavailable human renin inhibitors

New classes of de novo designed renin inhibitors are reported. Some of these compounds display excellent in vitro and in vivo activities toward human renin in a TGR model. The synthesis of these new types of mono- and bicyclic scaffolds are reported, and properties of selected compounds discussed.     

Drosophila cdk5 is needed for locomotive behavior and NMJ elaboration,but seems dispensable for synaptic transmission

Cyclin‐dependent kinase 5 (Cdk5) functions in postmitotic neuronal cells and play roles in cell differentiation, cell migration, axonal guidance, and synaptic function. Here, we demonstrate that Drosophila cdk5 is dispensable for adult viability and fertility, a feature that allows us to study its physiological function in the whole animal model. For the adult, cdk5 is needed for proper locomotion and flight performance. Larvae lacking cdk5 in the presynaptic tissue display abnormal crawling motion, and their neuromuscular junctions (NMJ) are elongated and contain a higher number of boutons that are smaller. As a result of these two counteracting effects, the total synaptic area/NMJ is similar to wild type, leading to normal synaptic transmission, indicating that a compensatory mechanism is capable of correcting the problem caused by the lack of cdk5. futsch, the Drosophila MAP1B homolog, is also involved in NMJ morphogenesis, and analysis of the NMJ phenotype of the double mutant futsch^K68; cdk5^? indicates that cdk5 is epistatic to futsch in this process. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

Role of the Lectin VIP36 in Post‐ER Quality Control of Human α1‐Antitrypsin

The leguminous‐type (L‐type) lectin VIP36 localizes to the Golgi apparatus and cycles early in the secretory pathway. In vitro, VIP36 binds high‐mannose glycans with a pH optimum of 6.5, a value similar to the luminal pH of the Golgi apparatus. Although the sugar‐binding properties of VIP36 in vitro have been characterized in detail, the function of VIP36 in the intact cell remains unclear as no convincing glycoprotein cargo has been identified. Here, we used yellow fluorescent protein (YFP) fragment complementation to identify luminal interaction partners of VIP36. By screening a human liver cDNA library, we identified the glycoprotein α1‐antitrypsin (α1‐AT) as a cargo of VIP36. The VIP36/α1‐AT complex localized to Golgi and endoplasmic reticulum (ER). In the living cell, VIP36 bound exclusively to the high‐mannose form of α1‐AT. The binding was increased when complex glycosylation was prevented by kifunensine and abolished when the glycosylation sites of α1‐AT were inactivated by mutagenesis. Silencing VIP36 accelerated α1‐AT transport, arguing against a role of VIP36 in anterograde traffic. The complex formed by VIP36 and α1‐AT in the Golgi recycled back to the ER. The combined data are most consistent with a function of VIP36 in post‐ER quality control of α1‐AT.