Ploidy level determination of Hedera (Araliaceae) with an emphasis on discussable species (Hedera hibernica)

Genome size is a helpful tool for circumscribing taxa at diverse taxonomic degrees (mostly species) and resolving intricate low-level taxonomies. The correct genome size in Hedera (Araliaceae) has long been discussed, and the ploidy levels of some taxa are still unclear. Twelve accessions of Hedera were measured via flow cytometry. Flow cytometry is a relatively rapid, inexpensive, and credible tool. Fresh leaves of Hedera samples and internal reference standard parsley (Petroselinum crispum) were stained with propidium iodide (PI). Flow cytometry measurements showed that for the accessions of 2CV (3.09 - 6.40 pg), the lowest amount of nuclear DNA was 3.09 pg for Hedera crebrescens (So), while the highest amount was 6.40 pg for H. hibernica “Hamilton,” representing a statistically significant difference. According to this study, the new taxon (H. crebrescens) is a diploid, though this taxon was previously considered H. hibernica (tetraploid).

Determination of Organosulfides from Onion Oil

Qualitative and semi-quantitative analysis of organosulfides extracted from oil obtained by steam distillation of yellow onions was performed by gas chromatography-mass spectrometry (GC-MS). The extraction efficiency of organosulfides from onion oil was evaluated across four solvents: dichloromethane; diethyl ether; n-pentane; and hexanes. Analysis of solvent extracted organosulfides by GC-MS provided qualitative results that support the use of dichloromethane over other solvents based on identification of 27 organosulfides from the dichloromethane extract as compared to 10 from diethyl ether; 19 from n-pentane; and 17 from hexanes. Semi-quantitative evaluation of organosulfides present in the dichloromethane extract was performed using diallyl disulfide as the internal reference standard. Three organosulfides were detected in the extract at ≥5 mg/kg; 18 organosulfides between 3–5 mg/kg; and six organosulfides at <3 mg/kg. The E/Z isomers of 1-propenyl propyl trisulfide were among the most prevalent components extracted from the onion oil across all solvents; and 3,6-diethyl-1,2,4,5-tetrathiane was among the most abundant organosulfides in all solvents except hexanes. The method described here for the extraction of organosulfides from steam distilled onion oil surveys common solvents to arrive at a qualitative and semi-quantitative method of analysis for agricultural products involving onions; onion oil; and secondary metabolites of Allium spp.

Histogram score contributes for reliability of DNA content estimatives in Brachiaria spp

Flow cytometry allows to estimate the DNA content of a large number of plants quickly. However, inadequate protocols can compromise the reliability of these estimates leading to variations in the values of DNA content the same species. The objective of this study was to propose an efficient protocol to estimate the DNA content of Brachiaria spp. genotypes with different ploidy levels using flow cytometry. We evaluated four genotypes (B. ruziziensis diploid and artificially tetraploidized; a tetraploid B. brizantha and a natural triploid hybrid), three buffer solutions (MgSO4, Galbraith and Tris-HCl) and three species as internal reference standards (Raphanus sativus, Solanum lycopersicum e Pisum sativum). The variables measured were: histogram score (1-5), coefficient of variation and estimation of DNA content. The best combination for the analysis of Brachiaria spp. DNA content was the use of MgSO4 buffer with R. sativus as a internal reference standard. Genome sizes expressed in picograms of DNA are presented for all genotypes and the importance of the histogram score on the results reliability of DNA content analyses were discussed.

DNA levels in somatic and germ line cells of a freshwater crustacean, Mesocyclops edax

Chromatin diminution--the fragmentation and elimination of chromosome Regions--provides an unusual opportunity to study genomic reorganization during development. Some species of copepods regularly excise and discard large amounts of nuclear DNA from presumptive somatic cell lines during early cleavage stages . To study this phenomenon in M. edax we determined DNA-Feulgen levels for more than 5,600 individual nuclei from squash preparations of 30 female and 25 male adults collected from lakes in Nova Scotia, Virginia and Florida. Fixation in 3:1 methanol/acetic acid was followed by squashing individual specimens in 45% acetic acid, freezing each slide in liquid N2 and thawing in absolute ethanol before air drying. Each series of slides was stained with the feulgen reaction for DNA and measured with a Vickers M86 scanning and integrating microdensitometer at 560 nm, using chicken RBC nuclei as an internal reference standard of 2.5 pg DNA per cell. This allowed us to ask several questions: are there differences in genome size (1) among specimens from different collecting localities, (2) between females and males at any single locality, and (3) between cells of germ line and somatic cell lineages?

Influence of Solution-Deposited Anions on Glow Discharge Relative Ion Yields

When standard addition methods were employed for quantification in glow discharge mass spectrometry (GDMS), relative ion yields for some, but not all, elements were observed to be greater when deposited as a solution residue than as a corresponding solid of the same composition. The origin of these effects was investigated and determined to result from the deposition of anions present in the solution standards. The enhancements could be duplicated from a solid by pretreatment with a solution containing the anion. The mechanism by which the enhancements occur is not well understood, but we believe it involves a form of reactive etching followed by collisional dissociation of the sputtered molecular species into metal ions. Preliminary research suggests that it may be possible to compensate for changes in relative ion yield with solution treatment by employing an appropriate internal reference standard, or to match closely the matrices of the sample and standard in such a manner that quantification by standard addition is possible.