ABSTRACT IMPACT: Characterizing the cellular composition of human adipose tissue may contribute to the prevention and/or treatment of obesity-associated metabolic diseases. OBJECTIVES/GOALS: Our aims in this study were to use single-cell techniques 1. to characterize cell types within the stromavascular fraction of human adipose tissue, 2. to identify subsets of cells within each type (sub-clustering), 3. to identify gene sets and pathways that may provide information on the function and significance of each cell cluster. METHODS/STUDY POPULATION: Abdominal subcutaneous adipose tissue samples from n=6 healthy volunteers (1M, 5F, age 28-38 y, BMI 24.5-63.0 kg/m2) were collected by aspiration or during surgery. In 3 subjects, all females, paired femoral samples were also collected. After collagenase digestion approximately n=10,000 cells/sample were used for single-cell RNA sequencing using the 10X Genomics platform. After QC and downstream analysis, data were analyzed in Seurat v.3.1.5. We identified first different cell types and then subclusters in an unbiased fashion. Gene Set Enrichment Analysis (GSEA) was used for pathway analysis. RESULTS/ANTICIPATED RESULTS: Progenitor markers are related to extracellular matrix, eg DCN (logFC progenitors vs other cells types=3.17, expressed in 99.7% (pct.1) of progenitors, 26.1% (pct.2) of others). Endothelial and pericytes shared markers like RBP7 (logFC=1.67, pct.1 0.88, pct.2 0.39); pericytes also showed unique markers, eg RGS5 (logFC=2.29, pct.1 0.89, pct.2 0.17). Progenitors are further divided into 11 sub-clusters, one of which showed enrichment of CD36 (high proliferation potential), FABP4 (differentiation), and of the novel marker PALMD (logFC=7.13, pct.1 0.94, pct.2 0.48). All p<10E-5. GSEA analysis suggests that inflammatory pathways are downregulated in both adipose progenitors and endothelial/pericyte cells in the femoral compared to the abdominal depot. DISCUSSION/SIGNIFICANCE OF FINDINGS: Single cell RNA sequencing provides unique insights into the molecular profile of cell types and the identification of novel subsets of cells within the human adipose tissue. Such cellular heterogeneity may explain differences in adipose function between individuals and eventually in the risk of obesity-associated metabolic diseases.

Raman scattered O VI features at 6825 Å and 7082 Å found in symbiotic stars are important spectroscopic tools to probe the mass transfer process. Adopting a Monte Carlo approach, we perform a profile analysis of Raman O VI features of the yellow SySt AG Draconis and make a comparison with the spectrum obtained with CFHT. It is assumed that the accretion flow is convergent on the entering side with enhanced O VI emission and the flux ratio F(1032)/F(1038)∼1, whereas on the opposite side the flow is divergent with low O VI emission and F(1032/F(1038)∼2. Our best fit to the spectrum is obtained from our model with a mass-loss rate of the giant ∼4 × 10−7 M⊙ yr−1. A slight red wing excess in the spectrum suggests the presence of bipolar neutral components receding in the directions perpendicular to the binary orbital plane with a speed ∼70km s−1

The genus Vicia L., one of the earliest domesticated plant genera, is a member of the legume tribe Fabeae of the subfamily Papilionoideae (Fabaceae). The taxonomic history of this genus is extensive and controversial, which has hindered the development of taxonomic procedures and made it difficult to identify and share these economically important crop resources. Species identification through DNA barcoding is a valuable taxonomic classification tool. In this study, four DNA barcodes (ITS2, matK, rbcL and psbA-trnH) were evaluated on 110 samples that represented 34 taxonomically best-known species in the Vicia genus. Topologies of the phylogenetic trees based on an individual locus were similar. Individual locus-based analyses could not discriminate closely related Vicia species. We proposed a concatenated data approach to increase the resolving power of ITS2. The DNA barcodes matK, psbA-trnH and rbcL were used as an additional tool for phylogenetic analysis. Among the four barcodes, three-barcode combinations that included psbA-trnH with any two of the other barcodes (ITS2, matK or rbcL) provided the best discrimination among Vicia species. Species discrimination was assessed with bootstrap values and considered successful only when all the conspecific individuals formed a single clade. Through sequencing of these barcodes from additional Vicia accessions, 17 of the 34 known Vicia species could be identified with varying levels of confidence. From our analyses, the combined barcoding markers are useful in the early diagnosis of targeted Vicia species and can provide essential baseline data for conservation strategies, as well as guidance in assembling germplasm collections.

Next-generation sequencing technique has been known as a useful tool for de novo transcriptome assembly, functional annotation of genes and identification of molecular markers. This study was carried out to mine molecular markers from de novo assembled transcriptomes of four chilli pepper varieties, the highly pungent ‘Saengryeg 211’ and non-pungent ‘Saengryeg 213’ and variably pigmented ‘Mandarin’ and ‘Blackcluster’. Pyrosequencing of the complementary DNA library resulted in 361,671, 274,269, 279,221, and 316,357 raw reads, which were assembled in 23,607, 19,894, 18,340 and 20,357 contigs, for the four varieties, respectively. Detailed sequence variant analysis identified numerous potential single-nucleotide polymorphisms (SNPs) and simple sequence repeats (SSRs) for all the varieties for which the primers were designed. The transcriptome information and SNP/SSR markers generated in this study provide valuable resources for high-density molecular genetic mapping in chilli pepper and Quantitative trait loci analysis related to fruit qualities. These markers for pepper will be highly valuable for marker-assisted breeding and other genetic studies.

The RDA Genebank at the National Agrobiodiversity Center (NAAS, RDA, Republic of Korea) has conserved about 182,000 accessions in 1777 species and is working at preserving agricultural genetic resources for the conservation and sustainable utilization of genetic diversity. The detection of genetic variability in conserved resources is important for germplasm management, but the molecular evaluation tools providing genetic information are insufficient for underutilized crops, unlike those for major crops. In this regard, the Korean National Agrobiodiversity Center has been developing microsatellite markers for several underutilized crops. We designed 3640 primer pairs flanking simple sequence repeat (SSR) motifs for 6310 SSR clones in 21 crop species. Polymorphic loci were revealed in each species (7–36), and the mean ratio of polymorphic loci to all the loci tested was 12%. The average allele number was 5.1 (2.8–10.3) and the expected heterozygosity 0.51 (0.31–0.74). Some SSRs were transferable to closely related species, such as within the genera Fagopyrum and Allium. These SSR markers might be used for studying the genetic diversity of conserved underutilized crops.

Distribution of wax in laser printer toner was observed using an ultra-high-voltage (UHV) and a medium-voltage transmission electron microscope (TEM). As the radius of the wax spans a hundred to greater than a thousand nanometers, its three-dimensional recognition via TEM requires large depth of focus (DOF) for a volumetric specimen. A tomogram with a series of the captured images would allow the determination of their spatial distribution. In this study, bright-field (BF) images acquired with UHV-TEM at a high tilt angle prevented the construction of the tomogram. Conversely, the Z-contrast images acquired by the medium-voltage TEM produced a successful tomogram. The spatial resolution for both is discussed, illustrating that the image degradation was primarily caused by beam divergence of the Z-contrast image and the combination of DOF and chromatic aberration of the BF image from the UHV-TEM.

Horizontal cylinders floating on liquid surfaces are mundanely observed, whose examples include the legs of aquatic arthropods and floating larvae, twigs and hairs. We study the force and energy required to lift the cylinder clear from the water surface, to evaluate the role of wettability, especially superhydrophobicity, in the adhesion of floating cylinders. We find that a drastic degree of energy saving is achieved when lifting a superhydrophobic cylinder as compared with a cylinder with moderate wettability. This can serve as a starting point to understand how the superhydrophobicity of the legs of water-walking insects help to propel them efficiently.

We investigated the solid phase crystallization (SPC) behavior of 1000 Å amorphous Si (a- Si) films deposited by plasma enhanced chemical vapor deposition (PECVD) at various temperatures and were able to enhance the grain size of the crystallized polysilicon films using double layers of a-Si filns. The deposition temperature of monolayer a-Si films varied from 200 to 400 °C and the films were recrystallized at 600 °C in nitrogen. As the deposition temperature increased, the incubation time was decreased and both the nucleation rate and growth rate were increased. Especially, the nucleation rate strongly depended on the deposition temperature.

Since the Si-SiO2 interface provides a large number of nucleation sites, it is desirable to suppress nucleation at the interface. As an idea we employed a structure with double layer a-Si films. The bottom a-Si layer deposited at lower temperature could suppress the nucleation at the Si-SiO2 interface while the top a-Si layer deposited at higher temperature could nucleate with a smaller number of nucleation sites. The incubation time and transformation behavior were determined by the deposition temperature of the top layer. As an example, the grain size of the double layer film deposited sequentially at 150 °C and 200 °C enhanced to 1.8 μm while that of the monolayer film deposited at 200 °C was 1.4 μm.