Nuclear DNA contents, rDNAs, and karyotype evolution in subgenus Vicia: III. The heterogeneous section Hypechusa.

Abstract: Nuclear DNA contents, automated karyotype analyses, and sequences of internal transcribed spacers from ribosomal genes have been determined in the species belonging to section Hypechusa of the sub-genus Vicia. Karyomorphological results and phylogenetic data generated from the comparison of rDNA ( genes coding for rRNA) sequences showed that sect. Hypechusa is not monophyletic; however, some monophyletic units are apparent ( one including Vicia galeata, V. hyrcanica, V. noeana, and V. tigridis, another including V. assyriaca, V. hybrida, V. melanops, V. mollis, and V. sericocarpa), which partly correspond to morphology-based infrasectional groups. The relationships among these species and the species in sections Faba, Narbonensis, Bithynicae, and Peregrinae have been also investigated. Nuclear DNA contents, automated karyotype analyses, and sequences of internal transcribed spacers from ribosomal genes have been determined in the species belonging to section Hypechusa of the subgenus Vicia. Karyomorphological results and phylogenetic data generated from the comparison of rDNA (genes coding for rRNA) sequences showed that sect. Hypechusa is not monophyletic; however, some monophyletic units are apparent (one including Vicia galeata, V. hyrcanica, V. noeana, and V. tigridis, another including V. assyriaca, V. hybrida, V. melanops, V. mollis, and V. sericocarpa), which partly correspond to morphology-based infrasectional groups. The relationships among these species and the species in sections Faba, Narbonensis, Bithynicae, and Peregrinae have been also investigated

The intergenic spacer region of the rDNA in Haplopappus gracilis (Nutt.) Gray

In this paper we provide further information on the genome organization of Haplopappus gracilis, one of the six angiosperms showing the lowest chromosome number, i.e. 2n=4, by determining the nucleotide sequence of the Intergenic Spacer region of the rRNA genes and its cytological localization on metaphase chromosomes. DNA sequence analysis reveals the occurring of a product of 4,382 bp in length, characterized by the presence of four blocks of different repeated sequences. Our analysis also evidenced putative promoter regions with three Transcription Initiation Sites for Polymerase I, as previously reported in Artemisia absinthium, belonging to the same Asteraceae family. A fluorescent in situ hybridization with the Intergenic Spacer probe indicates the presence of rDNA genes only in the satellited chromosomes of H. gracilis; besides differences in the signal intensity between homologous chromosomes were frequently observed, so suggesting, for these chromosome sites, the presence of a variable number of rDNA gene copies, even if a divergent chromatin organization in corresponding regions can not be ruled out

Characterization of normal and "albino" phenotypes in Erythrina crista-galli

We present here a characterization of Erythrina crista-galli L. (syn: Erythrina lamifolia Jacq.) seedlings, obtained from a plant from the Botanical Garden of Pisa University. This plant produces seeds that, during germination, have shown two different seedling phenotypes: normal (NT, 75%) and "albino" types (AT, 25%). Albino seedlings survive only 8–9 weeks and their growth is dramatically reduced when compared with wild type seedlings. Biochemical investigations have shown that albino seedlings completely lack chlorophyll and carotenoids and also soluble sugar levels are lower than in the normal type. We have also conducted sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) experiments and silver staining analysis on different protein extracts from shoots and leaves of both phenotypes, and demonstrated strong differences in protein patterns. The almost total absence of putative small and large RuBisCo bands in albino seedlings should be emphasized. We have also microspectrophotometrica..

Gitelman syndrome associated with chondrocalcinosis and severe neuropathy: a novel heterozygous mutation in SLC12A3 gene

Gitelman syndrome (GS) is an inherited salt-wasting tubulopathy characterized by hypocalciuria, hypokalemia, hypomagnesemia and metabolic alkalosis, due to inactivating mutations in the SLC12A3 gene. Symptoms may be systemic, neurological, cardiovascular, ophthalmological or musculoskeletal. We describe a 70 year-old patient affected by recurrent arthralgias, hypoesthesia and hyposthenia in all 4 limbs and severe hypokalemia, complicated by atrial flutter. Moreover, our patient reported eating large amounts of licorice, and was treated with medium-high dosages of furosemide, thus making diagnosis very challenging. Genetic analysis demonstrated a novel heterozygous mutation in the SLC12A3 gene; therefore, we diagnosed GS and started potassium and magnesium replacement. GS combined with chondrocalcinosis and neurological involvement is quite common, but this is the first case of an EMG-proven severe neuropathy associated with GS. Herein, we underline the close correlation between hypomagnesemia, chondrocalcinosis and neurological involvement. Moreover, we report a new heterozygous mutation in exon 23 (2738G>A), supporting evidence of a large genetic heterogeneity in this late-onset congenital tubulopathy

The elephant in the room of density functional theory calculations

Using multiwavelets, we have obtained total energies and corresponding atomization energies for the GGA-PBE and hybrid-PBE0 density functionals for a test set of 211 molecules with an unprecedented and guaranteed μHartree accuracy. These quasi-exact references allow us to quantify the accuracy of standard all-electron basis sets that are believed to be highly accurate for molecules, such as Gaussian-type orbitals (GTOs), all-electron numeric atom-centered orbitals (NAOs), and full-potential augmented plane wave (APW) methods. We show that NAOs are able to achieve the so-called chemical accuracy (1 kcal/mol) for the typical basis set sizes used in applications, for both total and atomization energies. For GTOs, a triple-ζquality basis has mean errors of ∼10 kcal/mol in total energies, while chemical accuracy is almost reached for a quintuple-ζbasis. Due to systematic error cancellations, atomization energy errors are reduced by almost an order of magnitude, placing chemical accuracy within reach also for medium to large GTO bases, albeit with significant outliers. In order to check the accuracy of the computed densities, we have also investigated the dipole moments, where in general only the largest NAO and GTO bases are able to yield errors below 0.01 D. The observed errors are similar across the different functionals considered here

molecular and chromosomal characterization of repeated and single copy dna sequences in the genome of dasypyrum villosum

Restriction fragment length polymorphism of ribosomal DNA repeated unit and single-copy DNA fragments and chromosomal distribution of a highly repeated sequence, have been studied to assess molecular markers and the extent of their heterogeneity in Dasypyrum villosum. Substantial variation has been found for the length of the intergenic spacer of ribosomal genes clustered in different alleles at Nor- VI locus of heterozygous individuals, but not within the cluster of rDNA of homozygous individuals. After Southern blots and hybridization to an intergenic spacer probe, each cluster of rDNA was detected as a single band with at least four variants differing for the number of 130 bp subrepeats in the intergenic spacer. One recombinant plasmid contained a 2270 bp DNA insert from the D. villosum genome that upon Sph I restriction endonuclease digestion was cleaved in three 380 bp repeat elements and one 1090 bp fragment. When Southern blots of Sph 1 digested D. villosum DNAs of different genotypes were hybridized to the 32P-labelled 380 bp repeat, a distinct ladder consisting of multiples of a basic repeat unit of about 380 bp in length was revealed on autoradiograms. The in situ hybridization of the 3H-labelled 380 bp repeat element showed that one chromosome pair (7V) was not labelled. In the other pairs, silver grains remained clustered at or near the telomeres. Dot-blot hybridization analysis of DNAs from a range of diploid, tetraploid, and hexaploid Triticeae species indicated that the 380 bp repeated element was a specific feature of the D. villosum genome. Other cloned DNA sequences of D. villosum showed a large restriction length polymorphism and one was located on V chromosomes

Anabolic effects and inhibition of interleukin 6 production induced by neridronate on human osteoblasts

Bisphosphonates (BPs) are pharmacological compounds widely used in the treatment of a variety of bone-related diseases, particularly where the bone-turnover is skewed in favour of osteolysis. The mechanisms by which BPs reduce bone-resorption directly acting on osteoclasts (OCs) are now largely clarified even at molecular level. The researches concerning the BPs effects on osteoblasts (OBs) have instead shown variable results. Objectives: We have investigated the efficacy of neridronate (NER), an amino-BP, as anabolic agent on human OBs. Moreover, we have tried to verify if NER is able to negatively modulate the production of IL-6 on OBs stimulated or not by the pro-inflammatory cytokine Il-1b. Methods: We have tested if different concentrations of NER (from 10-11M to 10-3M), added to primary human OB cultures, could affect the cells number, the endogenous cellular alkaline phosphatase (ALP) activity, the collagen I (COLI) synthesis, the formation of mineralized nodules and the IL-6 production. Our experimental approach was performed testing a wide range of NER concentrations because, under physiological conditions, OBs seems to be exposed to variable and transient levels of the drug. Results: Our results show that NER doesn't negatively affect in vitro the viability, proliferation and cellular activity of human OBs, even after 20 days of exposure to concentrations ²10-5 M (therapeutic dose). In addition, NER seems to enhance the differentiation of cultured OBs in mature bone-forming cells. A maximum increase of COL-I synthesis (+25% after 4 days; p<0.05), ALP activity (+50% after 10 days; p<0.01) and mineralized nodules (+48% after 20 days; p<0.05) was observed in cultures treated with NER 10-8M. A maximal reduction of IL-6 secretion (-24% on IL-1b stimulated cultures and -29% on unstimulated cultures) was observed for NER 10-9 M. Conclusions: These results encourage the use of neridronate in therapy of demineralizing metabolic bone disorders