Morphological analysis of female gametophyte development in the bel1 stk shp1 shp2 mutant

Abstract

In Arabidopsis thaliana, cell fate in developing ovules is determined by the action of the homeodomain factor BELL1 (BEL1) and of the MADS-box factors SEEDSTICK (STK), SHATTERPROOF1 (SHP1) and SHP2. The analysis of the bel1 and the stk shp1 shp2 mutants revealed that the functional megaspore is formed, however, it does not proceed into megagametogenesis. In the bel1 stk shp1 shp2, quadruple mutant megasporogenesis does not take place. In this article we describe a detailed morphological analysis of the quadruple mutant, and we discuss the possibility that BELL1, STK, SHP1 and SHP2 not only control integument identity determination and development, but that they might also play a role during megasporogenesis.     

Genetic diversity in the endangered Sicilian endemic Brassica rupestris: Proposals for a conservation strategy

Abstract

Brassica rupestris Raf. is a chasmophyte species that includes two subspecies, both endemic to Central-Western Sicily (Italy). Inter-Simple Sequence Repeat (ISSR) markers were used to detect genetic diversity within and among eight populations representative of the species' distribution range. High levels of genetic diversity were revealed both at the population (PPB = 53.88%, H _S = 0.212, Sh = 0.309) and at the species level (PPB = 96.55%, H _T = 0.307, Sh = 0.464). The correlation between genetic and geographical distances was negative (Mantel test, r = ?0.06, P < 0.95). The two subspecies of B. rupestris, subsp. rupestris and subsp. hispida, showed remarkable genetic similarity and molecular data did not unequivocally support their distinctness. The pattern of genetic variation revealed by our study bears important consequences for conservation management: It is desirable to preserve B. rupestris populations in situ with a “dynamic” strategy, while, ex situ conservation programmes might be improved to safeguard maximum genetic diversity.     

A nonradioisotope biomedical assay for intact oligonucleotide and its chain-shortened metabolites used for determination of exposure and elimination half-life of antisense drugs in tissue.

Rigorous extraction methods coupled with capillary gel electrophoresis (CGE) provide a basis for a nonradiolabel assay for quantitation of intact antisense drug and its numerous chain-shortened metabolites. As part of the validation of the CGE method, we compared the quantitation of unlabeled ISIS 3521 (ISI 641A) and its chain-shortened metabolites with total radioactivity of [(35)S]-ISIS 3521. ISIS 3521 was labeled on the fifth nucleotide linkage from the 5'-end with (35)S by well-established methods. Multiple tissues collected from rats after administration of [(35)S]-ISIS 3521 were assayed by both radiolabel (liquid scintillation spectroscopy) and CGE methods. The CGE method provided accurate quantitation of the drug and its metabolites in kidney cortex and liver tissues. The correlation between methods for multiple tissues over time was excellent with 88.5% of the measurements being statistically equivalent. These data suggest that CGE is an accurate means of quantitating oligonucleotide in tissue and that it compares favorably with traditional radiochemical techniques. Clearance half-lives for total measurable oligonucleotides were equivalent to clearance of total radioactivity in both liver and kidney with the longest clearance half-life associated with the kidney.     

The generic placement of a morphologically enigmatic species in Asteraceae: evidence from ITS sequences

 Bidens cordylocarpa is a high polyploid species restricted in distribution to stream sides in the mountains of Jalisco, Mexico. The morphologically enigmatic species was originally described as a member of the genus Coreopsis, but later transferred to Bidens, largely because the involucral bracts appear most similar to Bidens. Characters of the cypselae, often useful in generic placement, are of no value for this species because the fruits have features not detected in either Bidens or Coreopsis. Sequences from the internal transcribed spacer region of nuclear ribosomal DNA (ITS) were used to assess the relationships of Bidens cordylocarpa. The molecular phylogeny places B. cordylocarpa in a strongly supported clade of Mexican and South American Bidens, and provides more definitive evidence of relationships than morphology, chromosome number, or secondary chemistry. Molecular, morphological, and chromosomal data suggest that B. cordylocarpa is an ancient polyploid, perhaps the remnant of a polyploid complex. Received August 28, 2000 Accepted February 11, 2001     

Rapid, simple and sensitive high-performance liquid chromatographic method for detection and determination of acyclovir in human plasma and its use in bioavailability studies

A rapid, simple and sensitive reversed-phase high-performance liquid chromatographic (HPLC) method has been developed for the measurement of acyclovir concentrations in human plasma and its use in bioavailability studies is evaluated. Unchanged acyclovir has been quantified without the introduction of an internal standard using the present method. Human plasma proteins were selectively precipitated by the addition of 7% perchloric acid to spiked plasma samples or to the plasma samples obtained after acyclovir administration to human volunteers and the mixture was spun at 1000 g for 10 min. The supernatant was directly injected into a Novaflex C₁₈ column and detected at 254 nm. The mobile phase consisted of octane sulfonic acid buffer (pH 2.5) and methanol (92:08). The limit of quantitation for acyclovir in plasma was 20 ng/ml, which enabled the determination of the area under the curve (AUC) more precisely, that is, it is much closer to its extrapolated value. The present method has been successfully applied to samples from bioavailability studies.     

Kinetics of acetylation-deacetylation of angiotensin II. Intramolecular interactions of the tyrosine and histidine side-chains

The possible existence of intramolecular interactions involving the tyrosine and histidine residues in angiotensin II has been investigated by measuring the reactivities of the functional groups in the molecule. Angiotensin II catalyzed the hydrolysis of p-nitrophenylacetate in the pH range 6.6-8.2 at higher rates than were consistent with the reactivities of the free constituent functional groups, and had 2-4% of the activity of chymotrypsin between pH 6.6 and 7.5. Treatment of angiotensin II with acetic anhydride demonstrated that the tyrosine hydroxyl and the imidazole side-chain in angiotensin II acetylated and deacetylated at markedly higher rates than for the free amino acids, indicating increased nucleophilicities and the presence of intrinsic deacetylation mechanisms for these residues in angiotensin II. These findings are consistent with the presence of tyrosine hydroxyl-histidine-carboxylate charge relay system in ANG II in aqueous environments, and suggest that ANG II may act at membrane receptors by a mechanism which is analogous to that operating in serine proteases.     

Bloodmeal analysis in Culicoides midges collected near horses,donkeys and zebras in the Eastern Cape,South Africa

An upsurge in African horse sickness (AHS) in the Eastern Cape, South Africa, from 2006 led to an epidemiological reassessment of the disease there. Light trapping surveys carried out near horses, donkeys and zebras in 2014–2016 collected 39 species of Culicoides midge (Diptera: Ceratopogonidae) that are potential vectors of AHS. To establish if these midges fed on equids, DNA sequences were obtained from the gut contents of 52 female midges (35 freshly blood‐fed, 13 gravid and four parous), representing 11 species collected across 11 sites. Culicoides leucostictus fed on all three equids. Culicoides bolitinos, Culicoides imicola and Culicoides magnus fed on both horses and donkeys. Culicoides onderstepoortensis fed on donkeys, and Culicoides similis and Culicoides pycnostictus fed on zebras. Bloodmeals from cows, pigs, warthogs, impalas and a domestic dog were also identified in various species, but none of the midges tested had fed on birds. These results contribute to knowledge of the vectorial capacity of several species of Culicoides with regard to AHS in the Eastern Cape and point to potential reservoir hosts, of which donkeys, zebras and domestic dogs have previously been found to harbour AHS. Blood‐fed midges were also obtained throughout winter, indicating the potential for endemic AHS in the province.     

Action of pilocarpine on the normal frog heart and in pathology

Experiments on frogs were performed to examine the effect of the M-cholinomimetic pilocarpine on the heart. It was discovered that at concentrations of 10(-15)--10(-5) g/ml pilocarpine exerted only an adverse chronotropic effect on the perfused heart. When applied at a concentration of 10(-4) g/ml the drug produced a negative as well as a positive chronotropic effect. The latter occurred spasmodically (without progressive rise in the heart rate) in association with a slow heart rate. In some experiments such effects were preceded by a certain deceleration of the heart. In experiments with positive chronotropic effects, arrhythmias and sinoatrial dissociation were observed sometimes. Experiments with recording of the electrograms of the sinuses and lower parts showed that such effects were caused not by pacemaker acceleration but by the removal of the blockade of conduction, between the pacemaker and the atria. As far as the pacemaker is concerned, pilocarpine exerted only a negative chronotropic effect.