Transcription analysis of apple fruit development using cDNA microarrays

The knowledge of the molecular mechanisms underlying fruit quality traits is fundamental to devise efficient marker-assisted selection strategies and to improve apple breeding. In this study, cDNA microarray technology was used to identify genes whose expression changes during fruit development and maturation thus potentially involved in fruit quality traits. The expression profile of 1,536 transcripts was analysed by microarray hybridisation. A total of 177 genes resulted to be differentially expressed in at least one of the developmental stages considered. Gene ontology annotation was employed to univocally describe gene function, while cluster analysis allowed grouping genes according to their expression profile. An overview of the transcriptional changes and of the metabolic pathways involved in fruit development was obtained. As expected, August and September are the two months where the largest number of differentially expressed genes was observed. In particular, 85 genes resulted to be up-regulated in September. Even though most of the differentially expressed genes are involved in primary metabolism, several other interesting functions were detected and will be presented

Long walk to genomics : history and current approaches to genome sequencing and assembly

Genomes represent the starting point of genetic studies. Since the discovery of DNA structure, scientists have devoted great efforts to determine their sequence in an exact way. In this review we provide a comprehensive historical background of the improvements in DNA sequencing technologies that have accompanied the major milestones in genome sequencing and assembly, ranging from early sequencing methods to Next-Generation Sequencing platforms. We then focus on the advantages and challenges of the current technologies and approaches, collectively known as Third Generation Sequencing. As these technical advancements have been accompanied by progress in analytical methods, we also review the bioinformatic tools currently employed in de novo genome assembly, as well as some applications of Third Generation Sequencing technologies and high-quality reference genomes

Inheritance studies of apple scab resistance and identification of Rvi14, a new major gene that acts together with other broad-spectrum QTL

Scab, caused by the fungal pathogen Venturia inaequalis, is the most common disease of cultivated apple (Malus domestica). The fungal races 6 and 7 have now overcome the major resistance gene Vf, which is widely used in apple breeding programmes. New breeding strategies to achieve durable resistance are thus necessary. The aim of this study was to determine the genetic basis of quantitative resistance of the apple cultivar ‘Du¨lmener Rosenapfel’, known to be scab resistant under different environmental conditions. An F1 progeny derived from the cross between the susceptible cultivar ‘Gala’ and ‘Du¨lmener Rosenapfel’ was tested in a greenhouse with a multi-isolate inoculum of V. inaequalis. Rvi14, a new major gene that conditions a chlorotic-type reaction, was mapped on linkage group (LG) 6 in a genomic region not known to be involved in disease resistance. A further three quantitative trait loci (QTL) for resistance were identified. One co-localized with Rvi14 on LG6, whereas the remaining two were detected on LG11 and LG17, in genomic regions already reported to carry broad-spectrum QTL in other genetic backgrounds. Since a selective genotyping approach was used to detect QTL, an expectation-maximization (EM) computation was used to estimate the corrected QTL contributions to phenotypic variation and was validated by entire progeny genotyping

molecular enrichment for detection of s aureus in recreational waters

The identification of rapid methods for the control of recreational water and of aquatic environments with similar characteristics is necessary to provide adequate levels of health safety for users. Molecular techniques have been proposed in recent years as a viable alternative to traditional microbiological methods, as they offer various advantages and are less time consuming than traditional tests. An innovative protocol based on molecular enrichment that allows the identification of low concentrations of Staphylococcus aureus in recreational water has been developed. The method is based on the specific amplification of prokaryotic genomic DNA by the usage of universal primers for 23S rDNA; subsequently, a second amplification step is performed with specific real-time polymerase chain reaction (PCR) primers and probe. This approach shows sensitivity levels similar to those observed with microbiological tests, with the additional benefits of the specificity typical of nucleic acids techniques. This methodology is easily applicable also to other microbiological parameters, representing an important milestone in hygiene monitoring by the detection of specific pollution indicators

Molecular markers linked to the apple scab resistance gene Vbj derived from Malus baccata jackii

Breeding for scab-resistant apple cultivars by pyramiding several resistance genes in the same genetic background is a promising way to control apple scab caused by the fungus Venturia inaequalis. To achieve this goal, DNA markers linked to the genes of interest are required in order to select seedlings with the desired resistance allele combinations. For several apple scab resistance genes, molecular markers are already available; but until now, none existed for the apple scab resistance gene Vbj originating from the crab apple Malus baccata jackii. Using bulk segregant analysis, three RAPD markers linked to Vbj were first identified. These markers were transformed into more reliable sequence-characterised amplified region (SCAR) markers that proved to be co-dominant. In addition, three SSR markers and one SCAR were identified by comparing homologous linkage groups of existing genetic maps. Discarding plants showing genotype-phenotype incongruence (GPI plants) plants, a linkage map was calculated. Vbj mapped between the markers CH05e03 (SSR) and T6-SCAR, at 0.6cM from CH05e03 and at 3.9cM from T6-SCAR. Without the removal of the GPI plants, Vbj was placed 15cM away from the closest markers. Problems and pitfalls due to GPI plants and the consequences for mapping the resistance gene accurately are discussed. Finally, the usefulness of co-dominant markers for pedigree analysis is also demonstrate

Evolution of an insect immune barrier through horizontal gene transfer mediated by a parasitic wasp.

Genome sequencing data have recently demonstrated that eukaryote evolution has been remarkably influenced by the acquisition of a large number of genes by horizontal gene transfer (HGT) across different kingdoms. However, in depth-studies on the physiological traits conferred by these accidental DNA acquisitions are largely lacking. Here we elucidate the functional role of Sl gasmin, a gene of a symbiotic virus of a parasitic wasp that has been transferred to an ancestor of the moth species Spodoptera littoralis and domesticated. This gene is highly expressed in circulating immune cells (haemocytes) of larval stages, where its transcription is rapidly boosted by injection of microorganisms into the body cavity. RNAi silencing of Sl gasmin generates a phenotype characterized by a precocious suppression of phagocytic activity by haemocytes, which is rescued when these immune cells are incubated in plasma samples of control larvae, containing high levels of the encoded protein. Proteomic analysis demonstrates that the protein Sl gasmin is released by haemocytes into the haemolymph, where it opsonizes the invading bacteria to promote their phagocytosis, both in vitro and in vivo. Our results show that important physiological traits do not necessarily originate from evolution of pre-existing genes, but can be acquired by HGT events, through unique pathways of symbiotic evolution. These findings indicate that insects can paradoxically acquire selective advantages with the help of their natural enemies

Molecular markers linked to the apple scab resistance gene Vbj derived from Malus baccata jackii

Breeding for scab-resistant apple cultivars by pyramiding several resistance genes in the same genetic background is a promising way to control apple scab caused by the fungus Venturia inaequalis. To achieve this goal, DNA markers linked to the genes of interest are required in order to select seedlings with the desired resistance allele combinations. For several apple scab resistance genes, molecular markers are already available; but until now, none existed for the apple scab resistance gene Vbj originating from the crab apple Malus baccata jackii. Using bulk segregant analysis, three RAPD markers linked to Vbj were first identified. These markers were transformed into more reliable sequence-characterised amplified region (SCAR) markers that proved to be co-dominant. In addition, three SSR markers and one SCAR were identified by comparing homologous linkage groups of existing genetic maps. Discarding plants showing genotype\u2013phenotype incongruence (GPI plants) plants, a linkage map was calculated. Vbj mapped between the markers CH05e03 (SSR) and T6-SCAR, at 0.6 cM from CH05e03 and at 3.9 cM from T6-SCAR. Without the removal of the GPI plants, Vbj was placed 15 cM away from the closest markers. Problems and pitfalls due to GPI plants and the consequences for mapping the resistance gene accurately are discussed. Finally, the usefulness of co-dominant markers for pedigree analysis is also demonstrated

Antimicrobial Effectiveness of Innovative Photocatalysts: A Review

Waterborne pathogens represent one of the most widespread environmental concerns. Conventional disinfection methods, including chlorination and UV, pose several operational and environmental problems; namely, formation of potentially hazardous disinfection by-products (DBPs) and high energy consumption. Therefore, there is high demand for effective, low-cost disinfection treatments. Among advanced oxidation processes, the photocatalytic process, a form of green technology, is becoming increasingly attractive. A systematic review was carried out on the synthesis, characterization, toxicity, and antimicrobial performance of innovative engineered photocatalysts. In recent decades, various engineered photocatalysts have been developed to overcome the limits of conventional photocatalysts using different synthesis methods, and these are discussed together with the main parameters influencing the process behaviors. The potential environmental risks of engineered photocatalysts are also addressed, considering the toxicity effects presented in the literature

Microsatellite markers spanning the apple ( Malus x domestica Borkh.) genome

A new set of 148 apple microsatellite markers has been developed and mapped on the apple reference linkage map Fiesta x Discovery. One-hundred and seventeen markers were developed from genomic libraries enriched with the repeats GA, GT, AAG, AAC and ATC; 31 were developed from EST sequences. Markers derived from sequences containing dinucleotide repeats were generally more polymorphic than sequences containing trinucleotide repeats. Additional eight SSRs from published apple, pear, and Sorbus torminalis SSRs, whose position on the apple genome was unknown, have also been mapped. The transferability of SSRs across Maloideae species resulted in being efficient with 41% of the markers successfully transferred. For all 156 SSRs, the primer sequences, repeat type, map position, and quality of the amplification products are reported. Also presented are allele sizes, ranges, and number of SSRs found in a set of nine cultivars. All this information and those of the previous CH-SSR series can be searched at the apple SSR database ( http://www.hidras.unimi.it ) to which updates and comments can be added. A large number of apple ESTs containing SSR repeats are available and should be used for the development of new apple SSRs. The apple SSR database is also meant to become an international platform for coordinating this effort. The increased coverage of the apple genome with SSRs allowed the selection of a set of 86 reliable, highly polymorphic, and overall the apple genome well-scattered SSRs. These SSRs cover about 85% of the genome with an average distance of one marker per 15c