Identification of novel single nucleotide polymorphisms in the DGAT1 gene of buffaloes by PCR-SSCP

Diacylglycerol O-acyltransferase 1 (DGAT1) is a microsomal enzyme that catalyzes the final step of triglyceride synthesis. The DGAT1 gene is a strong functional candidate for determining milk fat content in cattle. In this work, we used PCR-SSCP (polymerase chain reaction-single-strand conformation polymorphism) and DNA sequencing to examine polymorphism in the region spanning exon 7 to exon 9 of the DGAT1 gene in Murrah and Pandharpuri buffaloes. Three alleles (A, B and C) and four novel single-nucleotide polymorphisms were identified in the buffalo DGAT1 gene. The frequencies of the alleles differed between the two buffalo breeds, with allele C being present in Murrah but not in Pandharpuri buffalo. The allele variation detected in this work may influence DGAT1 expression and function. The results described here could be useful in examining the association between the DGAT1 gene and milk traits in buffalo.     

Analysis of the mitochondrial genome of Schistocerca gregaria gregaria (Orthoptera: Acrididae)

In the present study, we present the full sequence of the mitochondrial genome of the African desert locust Schistocerca gregaria gregaria. The size of 15625 bp reported matches very well with mitochondrial genomes of other Orthopteriodea. The mitochondrial genome comprises 13 protein‐coding genes, two ribosomal RNAs and 22 t‐RNAs with two t‐RNA (trnD and trnK) rearrangements that are typical for the taxon Caelifera. We compared the sequence with 12 mitochondrial genes of Schistocerca gregaria flaviventris and Schistocerca americana and used some of these data to construct phylogenetic trees, which confirm the close relationship between the two subspecies S. g. flaviventris and S. g. gregaria. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 296–305.     

Dinoflagellate cyst biostratigraphy of the Opalinuston Formation (Middle Jurassic) in the Aalenian type area in southwest Germany and north Switzerland

Feist‐Burkhardt, S. & Pross, J. 2010: Dinoflagellate cyst biostratigraphy of the Opalinuston Formation (Middle Jurassic) in the Aalenian type area in southwest Germany and north Switzerland. Lethaia, Vol. 43, pp. 10–31. In order to provide a detailed dinoflagellate cyst stratigraphy of the Lower Aalenian Opalinuston Formation from the Aalenian type area, 68 samples from four boreholes and one outcrop section were analysed. The sample localities are Hausen an der Fils and Wittnau in southwest Germany, Weiach in north Switzerland and Mont Russelin in the Swiss Jura Mountains. Dinoflagellate cyst assemblages were recovered from the Late Toarcian Aalensis Zone to the Late Aalenian Murchisonae Zone. The samples yielded rich, well‐preserved and diverse assemblages with 51 dinoflagellate cyst taxa identified in total. The dinoflagellate cyst distribution data obtained from this study allow a high‐resolution biostratigraphical subdivision of the lowermost Middle Jurassic Opalinuston Formation into four palynostratigraphical units. First and last occurrences, acmes and consistent presence of the species Batiacasphaera sp. A, Evansia cf. granochagrinata, Kallosphaeridium praussii, Nannoceratopsis triangulata, Phallocysta? frommernensis and Wallodinium laganum were selected as the criteria for defining these units. The obtained high‐resolution palynostratigraphical scheme provides a basis for establishing and further refining early Middle Jurassic biostratigraphy in the Boreal and Tethyan realms. □Aalenian, biostratigraphy, dinoflagellate cysts, Germany, Jurassic, Switzerland, Toarcian.     

Vergleichende Untersuchungen zum Kontaktverhalten verschiedener Arten der Gattung Tilapia (Cichlidae,Pisces) und ihrer Bastarde

• 1 Tilapia tholloni (Substratbrüter), T. nilotica (weiblicher Maulbrüter) und T. heudeloti macrocephala (männlicher Maulbrüter) wurden künstlich erbrütet und ihre angeborenen Kontaktreaktionen in standardisierten Attrappenversuchen untersucht.
• 2 Das Kontaktverhalten muß während einer kritischen Phase (bei T. nilotica unter den angegebenen Versuchsbedingungen bis ungefähr zum 21. Tag nach dem Ablaichen) aktiviert werden, wenn es längere Zeit andauern soll (= Reaktionsphase). Die Reaktionsphase kann bei T. nilotica mehrere Wochen dauern. In ihr nimmt die Reaktionsstärke (Anzahl und Dauer der Kontakte) zunächst rasch zu, erreicht ein Maximum und nimmt dann allmählich wieder ab. Anstieg, Maximum und Abnahme sind an bestimmte Entwicklungsabschnitte gebunden, weitgehend unabhängig davon, ob die Tiere zuvor schon Kontaktverhalten geäußert haben oder nicht. Die Kontaktreaktionen unterscheiden sich u. a. durch die Dauer der Kontakte: tholloni = 0,7 Sek., nilotica = 86,5 Sek., heudeloti m. = 1,5 Sek. je Tag und Tier (Maximalwerte bei bestimmten, für alle Arten gleichen Versuchsbedingungen).
• 3 Von der Aufzuchttemperatur hängt es ab, in welchem Entwicklungsabschnitt die Reaktionsphase liegt. Das Reaktionsmaximum junger T. nilotica lag bei 24° C am 9., bei 29° C am Tag nach der Eiablage.
• 4 Geblendete nilotica-Jungfische zeigten nur zu Beginn der Reaktionsphase schwaches Kontaktverhalten, normale Tiere äußerten gegenüber einer durchsichtigen Glasattrappe abgeschwächtes, nur kurze Zeit dauerndes Kontaktverhalten. Das Kontaktverhalten wird durch mechanische (Strömung) und optische Reize ausgelöter und gesteuert.
• 5 Junge Maulbrüter aus kleinen Eiern erreichten eine längere Kontaktdauer als solche aus größeren. Nach künstlicher Reduktion der Dottermenge um 10–20%) erhöhte sich die Kontaktdauer bei jungen nilotica um 23,1%, die Zahl der Kontakte nahm gleichzeitig um 7,6% ab. Die ♀♀ der substratbrütenden T. mariae legen große Eier, die Jungen zeigten gegenüber Attrappen intensive Kontaktreaktionen. Es wird die Frage diskutiert, ob das Erreichen einer bestimmten Eigröße eine Voraussetzung für das Entstehen des Kontaktverhaltens gewesen sein könnte.
• 6 Die Kurzkontakte junger T. tholloni werden aus ihrer Orientierungsreaktion abgeleitet und als Vorstufe des Kontaktverhaltens gedeutet.
• 7 7. Mehrere Tilapia-Arten wurden künstlich gekreuzt. Bei Verwendung von tholloni-Sperma wiesen die Bastarde eine erhöhte Sterblichkeit auf. Aus der Kreuzung T. tholloni ♀ ～ T. nilotica ♂ (Substratbrüter ～ weiblicher Maulbrüter) gingen nur ♀♀ hervor. Die Kreuzung T. heudeloti macrocephala ～ T. nilotica (männlicher ～ weiblicher Maulbrüter) erbrachte fertile F₁- und F₂-Generationen sowie alle vier möglichen Rückkreuzungen. Bei der Vererbung des Kontaktverhaltens (gemessen an der Dauer der Kontakte) scheinen relativ wenig Erbfaktoren mitzuwirken, T. heudeloti m. erwies sich gegenüber nilotica als praevalent. Ein Teil der Bastarde aus der Kreuzung T. tholloni ♀ ～ T. nilotica ♂ (F₁) (Substratbrüter ～ weiblicher Maulbrüter) lag auf der Merkmalsskala zwischen den Ausgangsarten, der Rest verteilte sich sowohl auf den Bereich von tholloni als auch auf den Bereich von nilotica.

     

Formation and properties of [4Fe-4S] clusters on the IscU scaffold protein

Rapid and quantitative reductive coupling of two [2Fe-2S]2+ clusters to form a single [4Fe-4S]2+ cluster on the homodimeric IscU Fe-S cluster scaffold protein has been demonstrated by UV-visible absorption, M?ssbauer, and resonance Raman spectroscopies, using dithionite as the electron donor. Partial reductive coupling was also observed using reduced Isc ferredoxin, which raises the possibility that Isc ferredoxin is the physiological reductant. The results suggest that reductive coupling of adjacent [2Fe-2S]2+ clusters assembled on IscU provides a general mechanism for the final step in the biosynthesis of [4Fe-4S]2+ clusters. The [4Fe-4S]2+ center on IscU can be reduced to a S = 1/2[4Fe-4S]+ cluster (g parallel = 2.06 and g perpendicular = 1.92), but the low midpoint potential (< -570 mV) and instability of the reduced cluster argue against any physiological relevance for the reduced cluster. On exposure to O2, the [4Fe-4S]2+ cluster on IscU degrades via a semistable [2Fe-2S]2+ cluster with properties analogous to those of the [2Fe-2S]2+ center in [2Fe-2S]2+ IscU. It is suggested that the ability of IscU to accommodate either [2Fe-2S]2+ or [4Fe-4S]2+ clusters in response to cellular redox status and/or oxygen levels may provide an effective way to populate appropriately cluster-loaded forms of IscU for maturation of different types of [Fe-S] proteins.     

The two-domain structure of 5'-adenylylsulfate (APS) reductase from Enteromorpha intestinalis is a requirement for efficient APS reductase activity

5'-Adenylylsulfate (APS) reductase from Enteromorpha intestinalis (EiAPR) is composed of two domains that function together to reduce APS to sulfite. The carboxyl-terminal domain functions as a glutaredoxin that mediates the transfer of electrons from glutathione to the APS reduction site on the amino-terminal domain. To study the basis for the interdomain interaction, a heterologous system was constructed in which the C domain of EiAPR was fused to the carboxyl terminus of the APS reductase from Pseudomonas aeruginosa (PaAPR), an enzyme that normally uses thioredoxin as an electron donor and is incapable of using glutathione for this function. The hybrid enzyme, which retains the [4Fe-4S] cluster from PaAPR, was found to use both thioredoxin and glutathione as an electron donor for APS reduction. The ability to use glutathione was enhanced by the addition of Na2SO4 to the reaction buffer, a property that the hybrid enzyme shares with EiAPR. When the C domain was added as a separate component, it was much less efficient in conferring PaAPR with the ability to use glutathione as an electron donor, despite the fact that the separately expressed C domain functioned in two activities that are typical for glutaredoxins, hydroxyethyl disulfide reduction and electron donation to ribonucleotide reductase. These results suggest that the physical connection of the reductase and C domain on a single polypeptide is critical for the electron-transfer reaction. Moreover, the effect of Na2SO4 suggests that a water-ordering component of the reaction milieu is critical for the catalytic function of plant-type APS reductases by promoting the interdomain interaction.     

Modeling brewers' yeast flocculation

Flocculation of yeast cells occurs during the fermentation of beer. Partway through the fermentation the cells become flocculent and start to form flocs. If the environmental conditions, such as medium composition and fluid velocities in the tank, are optimal, the flocs will grow in size large enough to settle. After settling of the main part of the yeast the green beer is left, containing only a small amount of yeast necessary for rest conversions during the next process step, the lagering. The physical process of flocculation is a dynamic equilibrium of floc formation and floc breakup resulting in a bimodal size distribution containing single cells and flocs. The floc size distribution and the single cell amount were measured under the different conditions that occur during full scale fermentation. Influences on flocculation such as floc strength, specific power input, and total number of yeast cells in suspension were studied. A flocculation model was developed, and the measured data used for validation. Yeast floc formation can be described with the collision theory assuming a constant collision efficiency. The breakup of flocs appears to occur mainly via two mechanisms, the splitting of flocs and the erosion of yeast cells from the floc surface. The splitting rate determines the average floc size and the erosion rate determines the number of single cells. Regarding the size of the flocs with respect to the scale of turbulence, only the viscous subrange needs to be considered. With the model, the floc size distribution and the number of single cells can be predicted at a certain point during the fermentation. For this, the bond strength between the cells, the fractal dimension of the yeast, the specific power input in the tank and the number of yeast cells that are in suspension in the tank have to be known. Copyright 1998 John Wiley & Sons, Inc.     

[3H]Adenosine Transport in Synaptoneurosomes of Postmortem Human Brain

Abstract: Abstract: [³H]Adenosine transport was characterized in cerebral cortical synaptoneurosomes prepared from postmortem human brain using an inhibitor-stop/centrifugation method. The adenosine transport inhibitors dipyridamole and dilazep completely and rapidly blocked transmembrane fluxes of [³H]adenosine. For 5-s incubations, two kinetically distinguishable processes were identified, i.e., a high-affinity adenosine transport system with K_t and V_max values of 89 μM and 0.98 nmol/min/mg of protein, respectively, and a low-affinity adenosine transport system that did not appear to be saturable. For incubations with 1 μM [³H]adenosine as substrate, intrasynaptoneurosomal concentrations of [³H]adenosine were 0.26 μM at 5 s and 1 μM at 600 s. Metabolism of accumulated [³H]adenosine to adenine nucleotides was 15% for 5-s, 23% for 15-s, 34% for 30-s, 43% for 60-s, and 80% for 600-s incubations. The concentrations (μM) of total accumulated ³H-purines ([³H]-adenosine plus metabolites) at these times were 0.3, 0.5, 1.0, 1.3 and 5.6, respectively. These results indicate that in the presence of extensive metabolism, the intrasynaptoneurosomal accumulation of ³H-purines was higher than the initial concentration of 1 μM [³H]adenosine in the reaction medium. For 5-, 15-, 30-, 60-, and 600-s incubations in the presence of the adenosine deaminase inhibitor EHNA and the adenosine kinase inhibitor 5′-iodotubercidin, metabolism of the transported [³H]adenosine was 14, 14, 16, 14, and 38%, respectively. During these times, total ³H-purine accumulation was 0.3, 0.5, 0.5, 0.7, and 1.8 μM, respectively. Thus, the apparently “concentrative'’accumulation of ³H-purines can be prevented by inhibition of adenosine metabolism and, taken together, these results suggest that adenosine transport in at least synaptoneurosomes prepared from postmortem human brain is via a nonconcentrative and equilibrative system.     

Characterization of novel microsatellite loci for red clover (Trifolium pratense L.) from enriched genomic libraries

Twenty‐seven polymorphic microsatellite markers were isolated from red clover (Trifolium pratense). Allelic variability and cross‐species amplification were assessed on 24 red clover and eight white clover (Trifolium repens) genotypes. The number of alleles detected in red clover ranged from two to 25. Observed and expected heterozygosities were high with average values of 0.71 and 0.88, respectively. Five of the 27 loci were also successfully amplified from white clover, where two to 13 alleles were detected. These highly polymorphic microsatellite loci provide powerful tools for population genetic studies as well as for marker‐assisted selection in this important forage legume species.