ZigBee End Devices are not fully functional devices, they send information only to their parent node, which can be either a Router or Coordinator. Generally, End Devices form the bulk of sensor nodes in a network structure. In order to facilitate long battery life, ZigBee protocols allow for an easily programmable sleep mode for End Devices, periodically limiting computation and thus power consumption. Coordinator and Router nodes continually transmit beacon signals to alert child nodes of their presence. Upon waking, End Devices wait for a beacon signal and relay information before continuing their sleep cycle.There are two types of UART schemes available for Xbee modules, AT and API mode. AT mode is synonymous with transparent mode, each module has a single network destination and personal address. Configuration of network parameters must be done through command mode, either by the user or a micro controller. AT mode is useful for simple point-to-point communication or non-modular network topology. API mode is recommended for larger networks with more complicated overall topologies. Messages contain a personal and destination address, plastic flower buckets wholesale which can be changed during run time depending on message type, making mesh topology simple to implement.
Additionally, API mode contains packet delivery confirmation messages and has the option of escape parameters.Grapevine berry ripening can be divided into three major stages. In stage 1, berry size increases sigmoidally. Stage 2 is known as a lag phase where there is no increase in berry size. Stage 3 is considered the ripening stage. Veraison is at the beginning of the ripening stage and is characterized by the initiation of color development, softening of the berry and rapid accumulation of the hexoses, glucose and fructose. Berry growth is sigmoidal in Stage 3 and the berries double in size. Many of the flavor compounds and volatile aromas are derived from the skin and synthesized at the end of this stage. Many grape flavor compounds are produced as glycosylated, cysteinylated and glutathionylated precursors and phenolics and many of the precursors of the flavor compounds are converted to various flavors by yeast during the fermentation process of wine. Nevertheless, there are distinct fruit flavors and aromas that are produced and can be tasted in the fruit, many of which are derived from terpenoids, fatty acids and amino acids. Terpenes are important compounds for distinguishing important cultivar fruit characteristics. There are 69 putatively functional, 20 partial and 63 partial pseudogenes in the terpene synthase family that have been identified in the Pinot Noir reference genome. Terpene synthases are multi-functional enzymes using multiple substrates and producing multiple products.
More than half of the putatively functional terpene synthases in the Pinot Noir reference genome have been functionally annotated experimentally and distinct differences have been found in some of these enzymes amongst three grape varieties: Pinot Noir, Cabernet Sauvignon and Gewürztraminer. Other aromatic compounds also contribute significant cultivar characteristics. C13-norisoprenoids are flavor compounds derived from carotenoids by the action of the carotenoid cleavage dioxygenase enzymes . Cabernet Sauvignon, Sauvignon Blanc and Cabernet Franc are characterized by specific volatile thiols and methoxypyrazines. Enzymes involved in the production of these aromas have been recently characterized. Phenolic compounds play a central role in the physical mouthfeel properties of red wine; recent work relates quality with tannin levels. While the grape genotype has a tremendous impact on tannin content, the environment also plays a very large role in grape composition. The pathway for phenolic biosynthesis is well known, but the mechanisms of environmental influence are poorly understood. Ultimately, there is an interaction between molecular genetics and the environment. Flavor is influenced by climate, topography and viticultural practices. For example, water deficit alters gene expression of enzymes involved in aroma biosynthesis in grapes, which is genotype dependent, and may lead to increased levels of compounds, such as terpenes and hexyl acetate, that contribute to fruity volatile aromas. The grapevine berry can be subdivided into the skin, pulp and seeds. The skin includes the outer epidermis and inner hypodermis . A thick waxy cuticle covers the epidermis.
The hypodermal cells contain chloroplasts, which lose their chlorophyll at veraison and become modified plastids; they are the sites of terpenoid biosynthesis and carotenoid catabolism. Anthocyanins and tannins accumulate in the vacuoles of hypodermal cells. Pulp cells are the main contributors to the sugar and organic acid content of the berries. Pulp cells also have a much higher set of transcripts involved in carbohydrate metabolism, but a lower set of transcripts involved in lipid, amino acid, vitamin, nitrogen and sulfur metabolism than in the skins. Hormones can influence berry development and ripening. Concentrations of auxin, cytokinins and gibberellins tend to increase in early fruit development of the first stage. At veraison, these hormone concentrations have declined concomitant with a peak in abscisic acid concentration just before veraison. Auxin prolongs the Stage 2 lag phase and inhibits anthocyanin biosynthesis and color development in Stage 3. Grapevine, a non-climacteric fruit, is not very sensitive to ethylene; however, ethylene appears to be necessary for normal fruit ripening. Ethylene concentration is highest at anthesis, but declines to low levels upon fruit set; ethylene concentrations rise slightly thereafter and peak just before veraison then decline to low levels by maturity. Ethylene also plays a role in the ripening of another non-climacteric fruit, strawberry. ABA also appears to be important in grape berry ripening during veraison when ABA concentrations increase resulting in increased expression of anthocyanin biosynthetic genes and anthocyanin accumulation in the skin. ABA induces ABF2, a transcription factor that affects berry ripening by stimulating berry softening and phenylpropanoid accumulation. In addition, ABA affects sugar accumulation in ripening berries by stimulating acid invertase activity and the induction of sugar transporters. It is not clear whether ABA directly affects flavor volatiles , but there could be indirect effects due to competition for common precursors in the carotenoid pathway. Many grape berry ripening studies have focused on targeted sampling over a broad range of berry development stages, but generally with an emphasis around veraison, when berry ripening is considered to begin. In this study, a narrower focus is taken on the late ripening stages where many berry flavors are known to develop in the skin. We show that that the abundance of transcripts involved in ethylene signaling is increased along with those associated with terpenoid and fatty acid metabolism, particularly in the skin.Cabernet Sauvignon clusters were harvested in 2008 from a commercial vineyard in Paso Robles, California at various times after veraison with a focus on targeting °Brix levels near maturity. Dates and metabolic details that establish the developmental state of the berries at each harvest are presented in Additional file 1. Berries advanced by harvest date with the typical developmental changes for Cabernet Sauvignon: decreases in titratable acidity and 2- isobutyl-3-methoxypyrazine concentrations and increases in sugar and color . Transcriptomic analysis focused on four harvest dates having average cluster °Brix levels of 22.6, 23.2, 25.0 and 36.7. Wines made in an earlier study from grapes harvested at comparable levels of sugars or total soluble solids to those in the present study showed clear sensory differences. Six biological replicates, black flower buckets comprising two clusters each, were separated into skins and pulp in preparation for RNA extraction and transcriptomic analysis using the NimbleGen Grape Whole-Genome Microarray. Thus, a 4 × 2 factorial experimental design was established. A note of caution must be added here. There are high similarities amongst members in certain Vitis gene families , making it very likely that cross-hybridization can occur with probes on the microarray with high similarity to other genes.
We estimate approximately 13,000 genes have the potential for cross-hybridization, with at least one probe of a set of four unique probes for that gene on the microarray potentially cross-hybridizing with probes for another gene on the microarray. Genes with the potential for cross hybridization have been identified and are highlighted in light red in Additional file 2. The rationale to include them is that although individual genes can not be uniquely separated, the probe sets can identify a gene and its highly similar gene family members, thus, providing some useful information about the biological responses of the plant. An additional approach was taken, removing cross-hybridizing probes before quantitative data analysis . Many of the significant genes were unaffected by this processing, but 3600 genes were completely removed from the analysis. Thus, it was felt that valuable information was lost using such a stringent approach. The less stringent approach allowing for analysis of genes with potential cross hybridization was used here in the rest of the analyses. To assess the main processes affected by these treatments, the gene ontologies of significantly affected transcripts were analyzed for statistical significance using BinGO. Based on transcripts that had significant changes in abundance with °Brix level, 230 biological processes were significantly over represented in this group . The three top over represented processes were response to abiotic stress, bio-synthetic process, and response to chemical stimulus, a rather generic set of categories. Tissue differences were more revealing at the stage when flavors peak; 4865 transcripts that were significantly higher in skins compared to pulp at 23.2 °Brix were tested for over represented GO functional categories . Some of the top GO categories included photosynthesis, isoprenoid biosynthesis, and pigment biosynthesis . Some of the transcripts with the largest differences between skin and pulp at 23.2 °Brix are β-ketoacyl-CoA synthase , taxane 10-β-hydroxylase , wax synthase, a lipase, an ABC transporter, and phenylalanine ammonia-lyase . The abundance of 5716 transcripts was significantly higher in pulp than skin at 23.2 °Brix . Some of the top GO categories over represented were a variety of transport processes and small GTPase mediated signal transduction . Some of the transcripts with the largest differences in abundance with pulp greater than skin at 23.2 °Brix were polygalacturonase , flavonol synthase, stachyose synthase, an amino acid transporter, a potassium channel , and HRE2 . The transcript abundance of 2053 genes had significantly differential expression across °Brix levels and tissues . The top GO categories over represented in this set involved photosynthesis and phenylpropanoid metabolism, both associated with the berry skin . Other flavorcentric categories of the 57 categories over represented include aromatic compound biosynthesis, fatty acid metabolism and alcohol catabolism. This transcript set was further analyzed by dividing into 10 clusters using k-means clustering . The over represented GO categories were determined for each cluster . Eight of the 10 clusters had distinct over represented GO categories; two clusters did not have any over represented GO categories, meaning that the genes in these two clusters were assigned to GO categories of expected proportions when compared to the entire NimbleGen array. Clusters 1, 8, 9 and 10 had a large number of over represented categories. Many GO categories within a cluster are subsets of others in that cluster and were grouped together. For example, cluster 4 had four over represented GO categories, oxygen transport, gas transport, heat acclimation and response to heat. The four categories could be grouped into two, as two are subsets of the others; this is how they were listed in Table 1.To properly annotate the AP2/ERF super family of Vitis vinifera according to the IGGP Supernomenclature committee instructions, a phylogenetic tree was generated for the AP2/ERF super family of Arabidopsis thaliana and Vitis vinifera using the TAIR 10 and V1 gene models, respectively . The labeled family classifications were derived from the Arabidopsis naming scheme by Nakano et al.. There are 130 members in the VitisAP2/ERF super family in the Pinot Noir reference genome. However, the six paralogs of ERF6 discussed above belong to a Vitis vinifera clade in subfamily IX and are distinctly different or separate from any Arabidopsis subfamily IX ERF TFs . All of these TFs in this clade are orthologs of AtERF6. VviERF6L1 [UniProt: F6I2N8; VIT_16s0013g00900] had one of the most interesting profiles of the 12 members of this clade because its transcript abundance peaked at 23.2 °Brix . Using k-means clustering, VviERF6L1 fell within Cluster 8 with 369 transcripts, including five additional VviERF6 paralogs. The top GO categories associated with Cluster 8 were genes associated with terpenoid metabolism and pigment biosynthesis . Other interesting flavor associated categories included fatty acid and alcohol metabolism . Representative transcripts from Cluster 8 that were correlated with the transcript abundance profile of VviERF6L1 can be seen in Figure 4.