Fast Quantitative Analysis of timsTOF PASEF Data with MSFragger and IonQuant

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Highlights

• •MSFragger now supports raw timsTOF PASEF data.
• •IonQuant performs fast and accurate feature detection and quantification.
• •MSFragger and IonQuant provide excellent performance for timsTOF PASEF data.
• •Flexibility allows for complex analyses, such as semi-enzymatic and open search.

     

Insertion sequence analysis of protoplast fused strains of Lactococcus lactis ssp. cremoris

Abstract The use of insertion sequence probes for the analysis of fusants obtained following protoplast fusion is described. Hybridization of both total and plasmid DNA from parent and fusant strains with probes to IS 904 and IS S1 showed that of the four protoplast fusions examined, three appeared to involve a rearrangement of genetic material while in the fourth the fusant appeared similar to one of the parental strains. This method of analysis provides more information about the changes induced by protoplast fusion than that obtained by monitoring the acquisition or loss of individual characteristics.     

Flank spot number and its significance for systematics,taxonomy and conservation of the near‐threatened Mediterranean trout Salmo cettii: evidence from a genetically pure population

The number of spots on the flank of the Mediterranean trout Salmo cettii population from Tellesimo Stream in Sicily, Italy, varied between seven and 37, with a median and mode of 21, numbers much lower than those reported in the literature and different from those of other Italian trout populations. This finding could be ascribed to the phenotypic plasticity of the species or, alternatively, could provide evidence to support the different origin of Sicilian autochthonous trout. Given the ease of use of this character for the identification of native fish, it could be particularly important for conservation and management purposes.     

Colletotrichum fructicola and C. siamense are involved in chilli anthracnose in India

Chilli anthracnose is a major problem in India and worldwide. In this study, we investigated the phylogenetic relationships of 52 fungal isolates associated with chilli anthracnose in southern India. All the 52 isolates were sequenced for partial ITS/5.8S rRNA and glyceraldehyde-3-phosphate dehydrogenase (gapdh) genes and showed affinities with Colletotrichum siamense and C. fructicola within Colletotrichum. gloeosporioides species complex. Further, a reduced subset of 17 selected isolates was made and in a maximum parsimony analysis of a multigene data-set including partial ITS/5.8S rRNA, actin (act), calmodulin (cal), chitin synthase (chs1), gapdh and β-tubulin (tub2) gene sequence data, these fungal isolates clustered with the type strain of C. fructicola, except for strain MTCC 3439 that showed phylogenetic affinities with C. siamense. The pathogenicity tests involving two representative isolates: UASB-Cg-14 and MTCC 3439, confirmed the involvement of C. fructicola and C. siamense in the development of disease symptoms on fresh chilli fruits. This is the first report of the association of C. fructicola and C. siamense in causing chilli anthracnose in India.     

C-Terminal Protein Characterization by Mass Spectrometry using Combined Micro Scale Liquid and Solid-Phase Derivatization

A sample preparation method for protein C-terminal peptide isolation has been developed. In this strategy, protein carboxylate glycinamidation was preceded by carboxyamidomethylation and optional α- and ϵ-amine acetylation in a one-pot reaction, followed by tryptic digestion of the modified protein. The digest was adsorbed on ZipTip_C18 pipette tips for sequential peptide α- and ϵ-amine acetylation and 1-ethyl-(3-dimethylaminopropyl) carbodiimide-mediated carboxylate condensation with ethylenediamine. Amino group-functionalized peptides were scavenged on N-hydroxysuccinimide-activated agarose, leaving the C-terminal peptide in the flow-through fraction. The use of reversed-phase supports as a venue for peptide derivatization enabled facile optimization of the individual reaction steps for throughput and completeness of reaction. Reagents were exchanged directly on the support, eliminating sample transfer between the reaction steps. By this sequence of solid-phase reactions, the C-terminal peptide could be uniquely recognized in mass spectra of unfractionated digests of moderate complexity. The use of the sample preparation method was demonstrated with low-level amounts of a model protein. The C-terminal peptides were selectively retrieved from the affinity support and proved highly suitable for structural characterization by collisionally induced dissociation. The sample preparation method provides for robustness and simplicity of operation using standard equipment readily available in most biological laboratories and is expected to be readily expanded to gel-separated proteins.     

Automated identification of social interaction criteria in Drosophila melanogaster

The study of social behaviour within groups has relied on fixed definitions of an ‘interaction’. Criteria used in these definitions often involve a subjectively defined cut-off value for proximity, orientation and time (e.g. courtship, aggression and social interaction networks) and the same numerical values for these criteria are applied to all of the treatment groups within an experiment. One universal definition of an interaction could misidentify interactions within groups that differ in life histories, study treatments and/or genetic mutations. Here, we present an automated method for determining the values of interaction criteria using a pre-defined rule set rather than pre-defined values. We use this approach and show changing social behaviours in different manipulations of Drosophila melanogaster. We also show that chemosensory cues are an important modality of social spacing and interaction. This method will allow a more robust analysis of the properties of interacting groups, while helping us understand how specific groups regulate their social interaction space.     

Effectiveness of mobile apps in teaching field-based identification skills

It has been suggested that few students graduate with the skills required for many ecological careers, as field-based learning is said to be in decline in academic institutions. Here, we asked if mobile technology could improve field-based learning, using ability to identify birds as the study metric. We divided a class of ninety-one undergraduate students into two groups for field-based sessions where they were taught bird identification skills. The first group has access to a traditional identification book and the second group were provided with an identification app. We found no difference between the groups in the ability of students to identify birds after three field sessions. Furthermore, we found that students using the traditional book were significantly more likely to identify novel species. Therefore, we find no evidence that mobile technology improved students’ ability to retain what they experienced in the field; indeed, there is evidence that traditional field guides were more useful to students as they attempted to identify new species. Nevertheless, students felt positively about using their own smartphone devices for learning, highlighting that while apps did not lead to an improvement in bird identification ability, they gave greater accessibility to relevant information outside allocated teaching times.