Tumor Necrosis Factor Receptor SF10A (TNFRSF10A) SNPs Correlate With Corticosteroid Response in Duchenne Muscular Dystrophy

Background Duchenne muscular dystrophy (DMD) is a rare and severe X-linked muscular dystrophy in which the standard of care with variable outcome, also due to different drug response, is chronic off-label treatment with corticosteroids (CS). In order to search for SNP biomarkers for corticosteroid responsiveness, we genotyped variants across 205 DMD-related genes in patients with differential response to steroid treatment. Methods and Findings We enrolled a total of 228 DMD patients with identified dystrophin mutations, 78 of these patients have been under corticosteroid treatment for at least 5 years. DMD patients were defined as high responders (HR) if they had maintained the ability to walk after 15 years of age and low responders (LR) for those who had lost ambulation before the age of 10 despite corticosteroid therapy. Based on interactome mapping, we prioritized 205 genes and sequenced them in 21 DMD patients (discovery cohort or DiC = 21). We identified 43 SNPs that discriminate between HR and LR. Discriminant Analysis of Principal Components (DAPC) prioritized 2 response-associated SNPs in theTNFRSF10Agene. Validation of this genotype was done in two additional larger cohorts composed of 46 DMD patients on corticosteroid therapy (validation cohorts or VaC1), and 150 non ambulant DMD patients and never treated with corticosteroids (VaC2). SNP analysis in all validation cohorts (N= 207) showed that the CT haplotype is significantly associated with HR DMDs confirming the discovery results. Conclusion We have shown that TNFRSF10A CT haplotype correlates with corticosteroid response in DMD patients and propose it as an exploratory CS response biomarker

Harvest time and storage of “Soreli” kiwifruit (Actinidia chinensis Pl.)

Abstract “Soreli” is a yellow-fleshed tetraploid selection of Actinidia chinensis developed at the University of Udine (Italy) and released in 2008. It crops heavily, ripens early-mid October in the northern hemisphere, and produces large, single fruit with intense yellow flesh color. Because they mature early in the season, fruit are sometimes harvested prematurely and the low quality of these fruit threatens the development of the cultivar in the market. We report in the present paper a two-year trial on harvest time (at approx. 7.0 and 8.5 °Brix), the monitoring of ripening in cool store and the final quality of fruit. The second year a treatment with 1-MCP (1-methylcyclopropene) in the cool store was included in the experimental plan. Fruit were stored in the cool store with temperatures maintained at 0.0 0.5°C with ethylene removed. Ripening was monitored at two-weekly intervals until February of the following year. Considering the two harvest times, fruit of the first harvest were firmer, but their soluble solids concentration (SSC) at consumption time was 0.7 °Brix lower than fruit harvested later in both years. Ripening followed rather different patterns in the two years. Firmness decreased rapidly the first year, but in the second year did not fall below 4 kg cm-2 until 4 February when the trial was stopped. The first year the soluble solid content reached c. 14 °Brix when fully ripe, whereas in the second season fruit reached only 12 °Brix, probably because of a long period of rain in May and the beginning of June. Treatment with 1-MCP when fruit entered the refrigerated cell enhanced fruit storage, with differences more evident in fruit harvested later in the season. Soluble solids content, acidity and dry matter content were not influenced by the 1-MCP treatment. No significant difference among harvesting time and 1-MCP treatment was recorded for the other parameters analysed, such as core firmness and flesh color. Remarkably no fruit decay was found amongst any of the samples

SSR‐based DNA Fingerprinting of Fruit Crops

The DNA fingerprinting of fruit crops, based on DNA microsatellite markers which are considered to be the key markers for the molecular analysis of germplasm collections, is reviewed. Single sequence repeats (SSRs) remain the markers of choice for fingerprinting in humans, animals, plants, and other living organisms. This review, that considers 44 fruit species, provides a set of markers that are suitable for profiling accessions and that should make the databases produced in different laboratories more comparable. Every effort has been made to select SSR markers that are robust and easily scorable considering that such analyses are sometimes used in legal cases. The review first describes the basic protocols, procedures and methods of data analyses; it then describes the fingerprinting of individual species or groups of species, providing a set of SSR markers and appropriate guidance based on the revised literature and on the authors’ experience

Linkage map in kiwifruit: the choice of mapping populations

A genetic map of kiwifruit (Actinidia spp.) was constructed using microsatellite and AFLP markers and the pseudo-test-cross mapping strategy. Some 105 microsatellite loci, of the 251 isolated from A. chinensis, segregated in the interspecific cross A. chinensis x A. callosa in a 1:1 ratio or in a ratio that could mimic the classical backcross, and were mapped using 94 individuals of the progeny. AFLP markers were also produced using MseI and EcoRI restriction enzymes and 15 primer combinations. Two linkage maps were produced, one for each parent. Some 225 markers segregated from the female parent (86 SSR and 139 AFLP) and 168 markers/loci (35 SSR, 132 AFLP and the sex determinant) from the male parent. Disappointedly only a few SSR markers segregated from the male parent either because of lack of amplification (18%) or because of homozygosity (48%). As a result, the male map had large gaps and only 14 linkage groups were recognised has holologous in the two maps. A test of polymorphism was therefore carried out on four genotypes of A. chinensis (the SSR source species) and four species (A. callosa, A. eriantha, A. latifolia, and A. polygama) using 30 randomly selected SSRs markers. Some 25/30 SSR were successfully amplified in A. chinensis and 14 to 15 of them were heterozygous; 17 to 22 SSR were amplified in the remaining species and 4 to 11 of them were heterozygous. The analysis of allelic combination at the 25 SSR for all combinations of genotype pairs revealed that 79% of SSR could be mapped in A. chinensis x A. chinensis pairs, 66% in A. chinensis x other, and 45% in other x other, where \u2018other\u2019 means any species different from A. chinensis. A similar tendency (35% vs. 19% vs. 9%) was found in the percentage of anchor loci, which is loci that map in both parents and could be used to find homologous linkage groups. These results prove that an intra-specific cross between A. chinensis genotypes should allow mapping the highest number of SSRs and should yield the highest number of anchor markers. More general aspects dealing with the selection of parents for the pseudo-test-cross and linkage maps are discussed in the paper

Genotyping apple (Malus x domestica Borkh) heirloom germplasm collected and maintained by the Regional Administration of Friuli Venezia Giulia (Italy)

This paper reports the genetic diversity of apple germplasm collected in the Friuli Venezia Giulia region (Italy). The collection, maintained in three different locations, was represented by local varieties likely originated as chance seedlings admixed with varieties introduced since centuries from neighbour countries, for most of which the name and origin was lost with the time. A preliminary procedure described in the paper started from 469 molecular profiles analysed at 15 Simple Sequence Repeat (SSR) markers and allowed to identify the \u2018true-to-type\u2019 genotypes among those maintained in multiple locations. The set of the remaining 233 accessions was further reduced to 133 unique profiles by removing 100 synonymy. The flow cytometry analysis allowed to identify as much as 55 triploids (41.4%), whose state was confirmed by field observations on leaf size and the occurrence of triallelic profiles at several SSR markers. The remaining 78 diploid accessions were analysed for their genetic diversity, that is the number of alleles, observed and expected heterozygosity, polymorphism information content (PIC), the frequency of null alleles and the probability of identity for unrelated and full-sib genotypes. The paper provides a critical evaluation of the SSR markers adopted for the study, discusses the genetic diversity observed in the apple collection examined as well as the high occurrence of triploids compared with other apple collections described in the literature

Genetic Diversity of Walnut (Juglans Regia L.) in the Eastern Italian Alps

Juglans regia L. is distributed primarily across temperate and subtropical regions of the Northern Hemisphere. During the last glaciation, the species survived in refugial areas that in Europe included the Balkans and the Italian peninsula, two areas joined by a corridor represented by the Friuli Venezia Giulia region, where two germplasm reservoirs met and likely intercrossed during re-colonization after the last glaciation. In this work, two hundred and fifteen wild accessions native to the area were sampled, georeferenced, and genotyped with 20 microsatellite loci selected from the literature. The local accessions of this study displayed moderate genetic diversity with 80 alleles identified. The number of alleles/loci was 4.0 (4.7 alleles for the genomic SSRs (Simple Sequence Repeats) and 2.7 alleles per EST (Expressed Sequence Tag)-derived SSR, on average). An analysis of molecular variance (AMOVA) revealed that most of the molecular diversity was between individuals (nearly 98% of variation explained). The model-based clustering algorithms implemented either in STRUCTURE and GENELAND software revealed two clusters: The first one encompassed most of the samples and showed a great genetic admixture throughout the five sampling areas defined on the base of orographic characteristics of the region. The second cluster represented a small island with three samples traced back to an introduction from Russia at the beginning of the 20th century