Leaf rust (Puccinia triticina Eriks. & Henn.) is a major disease affecting durum wheat production. The Lr14a-resistant gene present in the durum wheat cv. Creso and its derivative cv. Colosseo is one of the best characterized leaf-rust resistance sources deployed in durum wheat breeding. Lr14a has been mapped close to the simple sequence repeat markers gwm146, gwm344 and wmc10 in the distal portion of the chromosome arm 7BL, a gene-dense region. The objectives of this study were: (1) to enrich the Lr14a region with single nucleotide polymorphisms (SNPs) and high-resolution melting (HRM)-based markers developed from conserved ortholog set (COS) genes and from sequenced Diversity Array Technology (DArT(®)) markers; (2) to further investigate the gene content and colinearity of this region with the Brachypodium and rice genomes. Ten new COS-SNP and five HRM markers were mapped within an 8.0 cM interval spanning Lr14a. Two HRM markers pinpointed the locus in an interval of <1.0 cM and eight COS-SNPs were mapped 2.1-4.1 cM distal to Lr14a. Each marker was tested for its capacity to predict the state of Lr14a alleles (in particular, Lr14-Creso associated to resistance) in a panel of durum wheat elite germplasm including 164 accessions. Two of the most informative markers were converted into KASPar(®) markers. Single assay markers ubw14 and wPt-4038-HRM designed for agarose gel electrophoresis/KASPar(®) assays and high-resolution melting analysis, respectively, as well as the double-marker combinations ubw14/ubw18, ubw14/ubw35 and wPt-4038-HRM-ubw35 will be useful for germplasm haplotyping and for molecular-assisted breeding.
In this study we investigate the combined effect on Heliothis virescens (Lepidoptera, Noctuidae) larvae of Aedes aegypti-Trypsin Modulating Oostatic Factor (Aea-TMOF), a peptide that inhibits trypsin synthesis by the gut, impairing insect digestive function, and Autographa californica nucleopolyhedrovirus Chitinase A (AcMNPV ChiA), an enzyme that is able to alter the permeability of the peritrophic membrane (PM). Aea-TMOF and AcMNPV ChiA were provided to the larvae by administering transgenic tobacco plants, co-expressing both molecules. Experimental larvae feeding on these plants, compared to those alimented on plants expressing only one of the two molecules considered, showed significantly stronger negative effects on growth rate, developmental time and mortality. The impact of AcMNPV ChiA on the PM of H. virescens larvae, measured as increased permeability to molecules, was evident after five days of feeding on transgenic plants expressing ChiA. This result was confirmed by in vitro treatment of PM with recombinant ChiA, extracted from the transgenic plants used for the feeding experiments. Collectively, these data indicate the occurrence of a positive interaction between the two transgenes concurrently expressed in the same plant. The hydrolytic activity of ChiA on the PM of tobacco budworm larvae enhances the permeation of TMOF molecules to the ectoperitrophic space, and its subsequent absorption. The permeation through the paracellular route of Aea-TMOF resulted in a spotted accumulation on the basolateral domain of enterocytes, which suggests the occurrence of a receptor on the gut side facing the haemocoel. The binding of the peptide, permeating at increased rates due to the ChiA activity, is considered responsible for the enhanced insecticide activity of the transgenic plants expressing both molecules. These data corroborate the idea that ChiA can be effectively used as gut permeation enhancer in oral delivery strategies of bioinsecticides targeting haemocoelic receptors. Dear Rene,We have revised the manuscript as requested. Below are the reviewers comments and for each issue raised our remedial action or an explanation to address their concerns is reported in bold.I think that this version is certainly improved, and we will be glad to consider any further suggestion you may have. All the best, FrancoReviewer #1: Fiandra et al.In this manuscript Fiandra et al present some interesting data on apparent enhancement of TMOF action by chitinase. Overall, this is a sound study that will be improved with the following modifications:1. What is the mechanism of action of TMOF? This should be introduced upfront. The action of TMOF in inhibiting trypsin synthesis and impeding larval digestive processes is key to the logic behind these experiments. Not until page 16 is this information provided for the reader! This information should actually be included in the abstract. Clearly introduce TMOF.The information requested has been added in the manuscript (Abstract and Introduction). Page 4What is the size o...
Gluten is a protein composite found in wheat and related grains including barley, rye, oat, and all their species and hybrids. Gluten matrix is a biomolecular network of gliadins and glutenins that contribute to the texture of pastries, breads, and pasta. Gliadins are mainly responsible for celiac disease, one of the most widespread food-related pathologies in Western world. In view of the importance of gliadin proteins, by combining the quartz crystal microbalance technology, a cheap and robust piezoelectric transducer, with the so-called photonic immobilization technique, an effective surface functionalization method that provides spatially oriented antibodies on gold substrates, we realized a sensitive and reliable biosensor for quantifying these analytes extracted from real samples in a very short time. The resulting immunosensor has a limit of detection of about 4 ppm and, more remarkably, shows excellent sensitivity in the range 7.5-15 ppm. This feature makes our device reliable and effective for practical applications since it is able to keep low the influence of false positives.
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