C5 Extract Induces Apoptosis in B16F10 Murine Melanoma Cells through Extrinsic and Intrinsic Apoptotic Pathways and Sub-G1 Phase Arrest

Purpose: To investigate the anti-cancer activities of C5 extract (C5E), a new herbal preparation from Korea, on B16F10 cells.Methods: The anti-proliferative effects of C5E were assessed by culturing B16F10 cells in the presence or absence of C5E. Cell cycle progression was analyzed by PI staining using flow cytometry. The quantities of apoptosis-inducing proteins were measured by Western blot.Results: C5E inhibited the proliferation of B16F10 cells but not human keratinocytes. C5E induced S phase arrest by interfering with cell regulatory factors such as cyclins B1, D1, D3, and E, and cyclindependent kinase 2, in B16F10 cells. Furthermore, immunoblot analysis confirmed that treatment with C5E induced apoptosis and cleaved caspase-3, poly (ADP-ribose) polymerase, via extrinsic pathway, whereas Bcl-2 expression was down-regulated. In addition, the suppression of cell proliferation by C5E is through down-regulation of p-Akt, up-regulation of phosphatase and tensin homolog protein expression via phosphoinositol 3 kinase survival signaling pathways in B16F10 cells. The combined cytotoxic effects of C5E and vinblastine generated 10 % increase in activity in contrast to the sum of the inhibitory effects of the individual agents.Conclusion: C5E shows promising anti-cancer activity and can be a useful adjuvant with vinblastine in combination therapeutic treatment of skin cancer.Keywords: Melanoma, Apoptosis, Anti-cancer, p53, Vinblastine, Cell cycle arrest, Caspas

Unassisted photoelectrochemical water splitting exceeding 7% solar to hydrogen conversion efficiency using photon recycling

Various tandem cell configurations have been reported for highly efficient and spontaneous hydrogen production from photoelectrochemical solar water splitting. However, there is a contradiction between two main requirements of a front photoelectrode in a tandem cell configuration, namely, high transparency and high photocurrent density. Here we demonstrate a simple yet highly effective method to overcome this contradiction by incorporating a hybrid conductive distributed Bragg reflector on the back side of the transparent conducting substrate for the front photoelectrochemical electrode, which functions as both an optical filter and a conductive counter-electrode of the rear dye-sensitized solar cell. The hybrid conductive distributed Bragg reflectors were designed to be transparent to the long-wavelength part of the incident solar spectrum (lambda>500 nm) for the rear solar cell, while reflecting the short-wavelength photons (lambda<500 nm) which can then be absorbed by the front photoelectrochemical electrode for enhanced photocurrent generation.114129Ysciescopu

Selection of a core set of RILs from Forrest × Williams 82 to develop a framework map in soybean

Soybean BAC-based physical maps provide a useful platform for gene and QTL map-based cloning, EST mapping, marker development, genome sequencing, and comparative genomic research. Soybean physical maps for “Forrest” and “Williams 82” representing the southern and northern US soybean germplasm base, respectively, have been constructed with different fingerprinting methods. These physical maps are complementary for coverage of gaps on the 20 soybean linkage groups. More than 5,000 genetic markers have been anchored onto the Williams 82 physical map, but only a limited number of markers have been anchored to the Forrest physical map. A mapping population of Forrest × Williams 82 made up of 1,025 F8 recombinant inbred lines (RILs) was used to construct a reference genetic map. A framework map with almost 1,000 genetic markers was constructed using a core set of these RILs. The core set of the population was evaluated with the theoretical population using equality, symmetry and representativeness tests. A high-resolution genetic map will allow integration and utilization of the physical maps to target QTL regions of interest, and to place a larger number of markers into a map in a more efficient way using a core set of RILs

Using Microsatellites to Understand the Physical Distribution of Recombination on Soybean Chromosomes

Soybean is a major crop that is an important source of oil and proteins. A number of genetic linkage maps have been developed in soybean. Specifically, hundreds of simple sequence repeat (SSR) markers have been developed and mapped. Recent sequencing of the soybean genome resulted in the generation of vast amounts of genetic information. The objectives of this investigation were to use SSR markers in developing a connection between genetic and physical maps and to determine the physical distribution of recombination on soybean chromosomes. A total of 2,188 SSRs were used for sequence-based physical localization on soybean chromosomes. Linkage information was used from different maps to create an integrated genetic map. Comparison of the integrated genetic linkage maps and sequence based physical maps revealed that the distal 25% of each chromosome was the most marker-dense, containing an average of 47.4% of the SSR markers and 50.2% of the genes. The proximal 25% of each chromosome contained only 7.4% of the markers and 6.7% of the genes. At the whole genome level, the marker density and gene density showed a high correlation (R2) of 0.64 and 0.83, respectively with the physical distance from the centromere. Recombination followed a similar pattern with comparisons indicating that recombination is high in telomeric regions, though the correlation between crossover frequency and distance from the centromeres is low (R2 = 0.21). Most of the centromeric regions were low in recombination. The crossover frequency for the entire soybean genome was 7.2%, with extremes much higher and lower than average. The number of recombination hotspots varied from 1 to 12 per chromosome. A high correlation of 0.83 between the distribution of SSR markers and genes suggested close association of SSRs with genes. The knowledge of distribution of recombination on chromosomes may be applied in characterizing and targeting genes

SNP assay to detect the ‘Hyuuga’ red-brown lesion resistance gene for Asian soybean rust

Asian soybean rust (ASR), caused by Phakopsora pachyrhizi Syd., has the potential to become a serious threat to soybean, Glycine max L. Merr., production in the USA. A novel rust resistance gene, Rpp?(Hyuuga), from the Japanese soybean cultivar Hyuuga has been identified and mapped to soybean chromosome 6 (Gm06). Our objectives were to fine-map the Rpp?(Hyuuga) gene and develop a high-throughput single nucleotide polymorphism (SNP) assay to detect this ASR resistance gene. The integration of recombination events from two different soybean populations and the ASR reaction data indicates that the Rpp?(Hyuuga) locus is located in a region of approximately 371 kb between STS70887 and STS70923 on chromosome Gm06. A set of 32 ancestral genotypes which is predicted to contain 95% of the alleles present in current elite North American breeding populations and the sources of the previously reported ASR resistance genes (Rpp1, Rpp2, Rpp3, Rpp4, Rpp5, and rpp5) were genotyped with five SNP markers. We developed a SimpleProbe assay based on melting curve analysis for SNP06-44058 which is tighly linked to the Rpp?(Hyuuga) gene. This SNP assay can differentiate plants/lines that are homozygous/homogeneous or heterozygous/heterogeneous for the resistant and susceptible alleles at the Rpp?(Hyuuga) locus

A pipeline for high throughput detection and mapping of SNPs from EST databases

Single nucleotide polymorphisms (SNPs) represent the most abundant type of genetic variation that can be used as molecular markers. The SNPs that are hidden in sequence databases can be unlocked using bioinformatic tools. For efficient application of these SNPs, the sequence set should be error-free as much as possible, targeting single loci and suitable for the SNP scoring platform of choice. We have developed a pipeline to effectively mine SNPs from public EST databases with or without quality information using QualitySNP software, select reliable SNP and prepare the loci for analysis on the Illumina GoldenGate genotyping platform. The applicability of the pipeline was demonstrated using publicly available potato EST data, genotyping individuals from two diploid mapping populations and subsequently mapping the SNP markers (putative genes) in both populations. Over 7000 reliable SNPs were identified that met the criteria for genotyping on the GoldenGate platform. Of the 384 SNPs on the SNP array approximately 12% dropped out. For the two potato mapping populations 165 and 185 SNPs segregating SNP loci could be mapped on the respective genetic maps, illustrating the effectiveness of our pipeline for SNP selection and validation

Single-nucleotide polymorphism discovery by high-throughput sequencing in sorghum

<p>Abstract</p> <p>Background</p> <p>Eight diverse sorghum (<it>Sorghum bicolor </it>L. Moench) accessions were subjected to short-read genome sequencing to characterize the distribution of single-nucleotide polymorphisms (SNPs). Two strategies were used for DNA library preparation. Missing SNP genotype data were imputed by local haplotype comparison. The effect of library type and genomic diversity on SNP discovery and imputation are evaluated.</p> <p>Results</p> <p>Alignment of eight genome equivalents (6 Gb) to the public reference genome revealed 283,000 SNPs at ≥82% confirmation probability. Sequencing from libraries constructed to limit sequencing to start at defined restriction sites led to genotyping 10-fold more SNPs in all 8 accessions, and correctly imputing 11% more missing data, than from semirandom libraries. The SNP yield advantage of the reduced-representation method was less than expected, since up to one fifth of reads started at noncanonical restriction sites and up to one third of restriction sites predicted <it>in silico </it>to yield unique alignments were not sampled at near-saturation. For imputation accuracy, the availability of a genomically similar accession in the germplasm panel was more important than panel size or sequencing coverage.</p> <p>Conclusions</p> <p>A sequence quantity of 3 million 50-base reads per accession using a <it>Bsr</it>FI library would conservatively provide satisfactory genotyping of 96,000 sorghum SNPs. For most reliable SNP-genotype imputation in shallowly sequenced genomes, germplasm panels should consist of pairs or groups of genomically similar entries. These results may help in designing strategies for economical genotyping-by-sequencing of large numbers of plant accessions.</p