Construction of cDNA libraries from trifluoroacetic acid-solvated amphibian skin secretions: molecular cloning of multiple bombinin-like peptide precursor transcripts from a library of yellow-bellied toad (Bombina variegata) secretion

Amphibian skin secretions are renowned as complex mixtures of bioactive peptides many of which are analogues of endogenous regulatory peptides. While skin secretions can be obtained non-invasively for peptidome analysis, parallel studies on the granular gland transcriptome required specimen sacrifice. The aim of the present study was to analyse archived skin secretions to determine the robustness of bioactive peptide precursor-encoding polyadenylated mRNAs in an attempt to extract maximum molecular information from rare samples. A range of solvated skin secretion samples were examined after lyophilisation for their potential to generate viable and comprehensive cDNA libraries based upon polyadenylated mRNA capture and amplification/cloning using appropriate commercial kits. Here we present unequivocal data that the granular gland transcriptome persists in a PCR amenable format even after storage for as long as 12 years in 0.1%(v/v) aqueous trifluoroacetic acid (TFA). We used a pooled skin secretion sample (2 ml) from the yellow-bellied toad, Bombina variegata (n = 14), containing the equivalent of 5 mg/ml of lyophilised skin secretion, that had been used in part for peptide isolation purposes in 1998 and had been stored at - 20 °C since that time. In the first cloning experiment, 12 different bombinin-like peptide precursor cDNAs were cloned encoding 17 different bombinins, the majority of which were novel. Subsequently, bombesin and bradykinin-related peptide precursor transcripts have been cloned successfully. These data illustrate the unexpected stability/longevity of the transcriptome in these secretions — a finding with implications for both this field of research and for the wider field of molecular biology.

Kinematics of the ring-like nebula SuWt 2

We present the first detailed spatio-kinematical analysis and modelling of the southern planetary nebula SuWt 2. This object presents a problem for current theories of planetary nebula formation and evolution, as it is not known to contain a central post-main-sequence star. Deep narrow-band [NII]6584Å images reveal the presence of faint bipolar lobes emanating from the edges of the nebular ring. Long-slit observations of the Ha and [NII]6584Å emission lines were obtained using the ESO (European Southern Observatory) Multi-Mode Instrument on the 3.6-m ESO New Technology Telescope. The spectra reveal the nebular morphology as a bright torus encircling the waist of an extended bipolar structure. By deprojection, the inclination of the ring is found to be 68° +/- 2° (cf. ~90° for the double A-type binary believed to lie at the centre of the nebula), and the ring expansion velocity is found to be 28 kms-1. Our findings are discussed with relation to possible formation scenarios for SuWt 2. Through comparison of the nebular heliocentric systemic velocity, found here to be -25 +/- 5km s-1, and the heliocentric systemic velocity of the double A-type binary, we conclude that neither component of the binary could have been the nebular progenitor. However, we are unable to rule out the presence of a third component to the system, which would have been the nebula progenitor.

Dielectronic recombination in He-like, Li-like, and Be-like highly charged ions in the KLL and KLM manifolds

This paper reports a systematic study of the dependence on atomic number of the dielectronic recombination resonance strengths for He-like, Li-like and Be-like ions. Recent measurements of dielectronic recombination resonance strengths for the KLL and KLM manifolds for iron, yttrium, iodine, holmium, and bismuth are also described. The resonance strengths were normalized to calculated electron impact ionization cross sections. The measured resonance strengths generally agree well with theoretical calculations using the distorted wave approximation. However, KLM resonance strength measurements on high atomic number open-shell ions gave higher values than those suggested by calculations. Using recently measured data, along with existing results, scaling laws have been generated as a function of atomic number for He-like, Li-like, and Be-like ions in the KLL and KLM manifolds.