A mutation in amino acid permease AAP6 reduces the amino acid content of the Arabidopsis sieve elements but leaves aphid herbivores unaffected

The aim of this study was to investigate the role of the amino acid permease gene AAP6 in regulating phloem amino acid composition and then to determine the effects of this altered diet on aphid performance. A genotype of Arabidopsis thaliana (L.) was produced in which the function of the amino acid permease gene AAP6 (At5g49630) was abolished. Plants homozygous for the insertionally inactivated AAP6 gene had a significantly larger mean rosette width than the wild type and a greater number of cauline leaves. Seeds from the aap6 mutant were also significantly larger than those from the wild-type plants. Sieve element (SE) sap was collected by aphid stylectomy and the amino acids derivatized, separated, and quantified using Capillary Electrophoresis with Laser Induced Fluorescence (CE-LIF). In spite of the large variation across samples, the total amino acid concentration of SE sap of the aap6 mutant plants was significantly lower than that of the wild-type plants. The concentrations of lysine, phenylalanine, leucine, and aspartic acid were all significantly lower in concentration in the aap6 mutant plants compared with wild-type plants. This is the first direct demonstration of a physiological role for an amino acid transporter in regulating SE composition in vivo. The amino acid availability in sieve element sap is thought to be the major limiting factor for aphid growth and reproduction. Despite the changes in their diet, the aphid Myzus persicae (Sulzer) displayed only small changes in feeding behaviour on mutant plants when measured using the Electronic Penetration Graph (EPG) technique. Salivation by the aphid into the SE (E1 phase) was increased on mutant plants but there was no significant effect on other feeding EPG behaviours, or in the rate of honeydew production. Consistent with the small effect on aphid feeding behaviour, there was only a small effect of reduced sieve element amino acid concentration on aphid reproduction. The data are discussed in relation to the regulation of phloem composition and the role of phloem amino acids in regulating aphid performance

Genetic Dissection of Complex Fruit Quantitative Traits in Peach Progen

Major research efforts in peach are dedicated to the discovery of genomic variants causing phenotypic effects in complex fruit traits such as: maturity date (MD), fruit size (FW), sugar (SSC) and acid content (TA), flesh texture (slow softening, SSf) and resistance to brown rot by Monilinia spp. (BRr). Five segregating progenies showing phenotypic variation for at least one of these traits are available in our experimental fields. For SSC and TA, an already validated approach based on Near-InfraRed spectroscopy (NIR), is being applied to phenotype some segregating progenies. For SSf and BRr instead, trait characterization has been performed, resulting in the identification of co-factor traits and definition of standardized phenotyping tools, which are currently applied in the characterization of segregating material (in the context of FruitBreedomics EU project). High-density linkage maps have been constructed with genotypic data obtained from IPSC Illumina 9K SNP chip (Italian Drupomics and FruitBreedomics frameworks) and Genotyping-by-Sequencing (GBS). Additionally, parents of these progenies have been re-sequenced (30-40x) and genetic variants present along their genomes have been identified. Multiple-QTL models (MQM) coupled with the use of co-factor traits is leading to the discovery of significant QTLs. Genomic variants are explored within QTL intervals on the genomes of progeny parents, in order to identify possible mutations causing phenotypic differences, and develop markers for marker-assisted selection approaches

Evaluation of Variability in the Sweet Orange Germplasm through Next Generation Clonal Fingerprinting

The great phenotypic variability characterizing the sweet orange [Citrus sinensis(L.) Osbeck] germplasm arises from spontaneous bud mutations, causing a diversification into major groups (common, Navel and blood oranges). A huge divergence also occurred within each varietal group. The genetic basis of such variability, also including nutritional and qualitative traits (ripening time, colour, fruit shape, acidity, sugars), is currently uncharacterized, and therefore not exploitable. With the aim of describing the somatic mutation events in the sweet orange group a deep-sequencing of 20 Italian and foreign accessions was performed by Illumina platform, allowing the identification of single nucleotide polymorphisms (SNPs), structural variants (SVs) and large deletions, specific to each varietal group or clone-specific. A subset of SNPs used for the design of two 384 SNP - GoldenGate Assays allowed to genotype 225 CREA sweet orange accessions. The developed markers represent the first reliable molecular tools able to unambiguously fingerprint each somatic mutant. Moreover, they might be used to associate mutations with phenotypic traits, and are a powerful tool for traceability. By using the GoldenGate assay, we have been able to fingerprint several blood orange clones starting from DNAs isolated from leaves or juice. These tools will potentially provide the consumer with a guarantee on the quality and origin of juices, avoiding eventual frauds

A Mutation in Amino Acid Permease AAP6 Reduces the Amino Acid Content of the Arabidopsis Sieve Elements but Leaves Aphid Herbivores Unaffected.

The aim of this study was to investigate the role of the amino acid permease gene AAP6 in regulating phloem amino acid composition and then to determine the effects of this altered diet on aphid performance. A genotype of Arabidopsis thaliana (L.) was produced in which the function of the amino acid permease gene AAP6 (At5g49630) was abolished. Plants homozygous for the insertionally inactivated AAP6 gene had a significantly larger mean rosette width than the wild type and a greater number of cauline leaves. Seeds from the aap6 mutant were also significantly larger than those from the wild-type plants. Sieve element (SE) sap was collected by aphid stylectomy and the amino acids derivatized, separated, and quantified using Capillary Electrophoresis with Laser Induced Fluorescence (CE-LIF). In spite of the large variation across samples, the total amino acid concentration of SE sap of the aap6 mutant plants was significantly lower than that of the wild-type plants. The concentrations of lysine, phenylalanine, leucine, and aspartic acid were all significantly lower in concentration in the aap6 mutant plants compared with wild-type plants. This is the first direct demonstration of a physiological role for an amino acid transporter in regulating SE composition in vivo. The amino acid availability in sieve element sap is thought to be the major limiting factor for aphid growth and reproduction. Despite the changes in their diet, the aphid Myzus persicae(Sulzer) displayed only small changes in feeding behaviour on mutant plants when measured using the Electronic Penetration Graph (EPG) technique. Salivation by the aphid into the SE (E1 phase) was increased on mutant plants but there was no significant effect on other feeding EPG behaviours, or in the rate of honeydew production. Consistent with the small effect on aphid feeding behaviour, there was only a small effect of reduced sieve element amino acid concentration on aphid reproduction. The data are discussed in relation to the regulation of phloem composition and the role of phloem amino acids in regulating aphid performance

Deletion of the miR172 target site in a TOE-type gene is a strong candidate variant for dominant double-flower trait in Rosaceae

Double flowers with supernumerary petals have been selected by humans for their attractive appearance and commercial value in several ornamental plants, including Prunus persica (peach), a recognized model for Rosaceae genetics and genomics. Despite the relevance of this trait, knowledge of the underlying genes is limited. Of two distinct loci controlling the double-flower phenotype in peach, we focused on the dominant Di2 locus. High-resolution linkage mapping in five segregating progenies delimited Di2 to an interval spanning 150858bp and 22 genes, including Prupe.6G242400 encoding an euAP2 transcription factor. Analyzing genomic resequencing data from single- and double-flower accessions, we identified a deletion spanning the binding site for miR172 in Prupe.6G242400 as a candidate variant for the double-flower trait, and we showed transcript expression for both wild-type and deleted alleles. Consistent with the proposed role in controlling petal number, Prupe.6G242400 is expressed in buds at critical times for floral development. The indelDi2 molecular marker designed on this sequence variant co-segregated with the phenotype in 621 progenies, accounting for the dominant inheritance of the Di2 locus. Further corroborating the results in peach, we identified a distinct but similar mutation in the ortholog of Prupe.6G242400 in double-flower roses. Phylogenetic analysis showed that these two genes belong to a TARGET OF EAT (TOE)-type clade not represented in Arabidopsis, indicating a divergence of gene functions between AP2-type and TOE-type factors in Arabidopsis and other species. The identification of orthologous candidate genes for the double-flower phenotype in two important Rosaceae species provides valuable information to understand the genetic control of this trait in other major ornamental plants. Significance Statement We used peach as a model to gain insight into the molecular basis of double flowers, an important trait in many ornamental plants. We propose that a deletion causes a TOE-type transcription factor to escape miR172-mediated repression, in turn resulting in an increased number of petals, as corroborated by results on the orthologous gene in rose

A new intra-specific and high-resolution genetic map of eggplant based on a ril population, and location of QTLS related to plant anthocyanin pigmentation and seed vigour

Eggplant is the second most important solanaceous berry-producing crop after tomato. Despite mapping studies based on bi-parental progenies and GWAS approaches having been performed, an eggplant intraspecific high-resolution map is still lacking. We developed a RIL population from the intraspecific cross ‘305E40’, (androgenetic introgressed line carrying the locus Rfo-Sa1 conferring Fusarium resistance) x ‘67/3’ (breeding line whose genome sequence was recently released). One hundred and sixty-three RILs were genotyped by a genotype-by-sequencing (GBS) approach, which allowed us to identify 10,361 polymorphic sites. Overall, 267 Gb of sequencing data were generated and ~773 M Illumina paired end (PE) reads were mapped against the reference sequence. A new linkage map was developed, including 7249 SNPs assigned to the 12 chromosomes and spanning 2169.23 cM, with iaci@liberoan average distance of 0.4 cM between adjacent markers. This was used to elucidate the genetic bases of seven traits related to anthocyanin content in different organs recorded in three locations as well as seed vigor. Overall, from 7 to 17 QTLs (at least one major QTL) were identified for each trait. These results demonstrate that our newly developed map supplies valuable information for QTL fine mapping, candidate gene identification, and the development of molecular markers for marker assisted selection (MAS) of favorable alleles

Integrative genomics approaches validate PpYUC11-like as candidate gene for the stony hard trait in peach (P. persica L. Batsch)

Texture is one of the most important fruit quality attributes. In peach, stony hard (SH) is a recessive monogenic trait (hd/hd) that confers exceptionally prolonged firm flesh to fully ripe fruit. Previous studies have shown that the SH mutation affects the fruit ability to synthesize appropriate amounts of indol-3-acetic acid (IAA), which orchestrates the ripening processes through the activation of system 2 ethylene pathway. Allelic variation in a TC microsatellite located within the first intron of PpYUC11-like (a YUCCA-like auxin-biosynthesis gene) has been recently proposed as the causal mutation of the SH phenotype

PeachVar-DB : Curated Collection of Genetic Variations for the Interactive Analysis of Peach Genome Data

Applying next-generation sequencing (NGS) technologies to species of agricultural interest has the potential to accelerate the understanding and exploration of genetic resources. The storage, availability and maintenance of huge quantities of NGS-generated data remains a major challenge. The PeachVar-DB portal, available at http://hpc-bioinformatics.cineca.it/peach, is an open-source catalog of genetic variants present in peach (Prunus persica L. Batsch) and wild-related species of Prunus genera, annotated from 146 samples publicly released on the Sequence Read Archive (SRA). We designed a user-friendly web-based interface of the database, providing search tools to retrieve single nucleotide polymorphism (SNP) and InDel variants, along with useful statistics and information. PeachVar-DB results are linked to the Genome Database for Rosaceae (GDR) and the Phytozome database to allow easy access to other external useful plant-oriented resources. In order to extend the genetic diversity covered by the PeachVar-DB further, and to allow increasingly powerful comparative analysis, we will progressively integrate newly released data