Kyungho Won

The Draft Genome Sequence of European Pear (Pyrus communis L. 'Bartlett')

We present a draft assembly of the genome of European pear (Pyrus communis) ‘Bartlett’. Our assembly was developed employing second generation sequencing technology (Roche 454), from single-end, 2 kb, and 7 kb insert paired-end reads using Newbler (version 2.7). It contains 142,083 scaffolds greater than 499 bases (maximum scaffold length of 1.2 Mb) and covers a total of 577.3 Mb, representing most of the expected 600 Mb Pyrus genome. A total of 829,823 putative single nucleotide polymorphisms (SNPs) were detected using re-sequencing of ‘Louise Bonne de Jersey’ and ‘Old Home’. A total of 2,279 genetically mapped SNP markers anchor 171 Mb of the assembled genome. Ab initio gene prediction combined with prediction based on homology searching detected 43,419 putative gene models. Of these, 1219 proteins (556 clusters) are unique to European pear compared to 12 other sequenced plant genomes. Analysis of the expansin gene family provided an example of the quality of the gene prediction and an insight into the relationships among one class of cell wall related genes that control fruit softening in both European pear and apple (Malus×domestica). The ‘Bartlett’ genome assembly v1.0 (http://www.rosaceae.org/species/pyrus/pyrus_communis/genome_v1.0) is an invaluable tool for identifying the genetic control of key horticultural traits in pear and will enable the wide application of marker-assisted and genomic selection that will enhance the speed and efficiency of pear cultivar development.

Identification of Pyrus single nucleotide polymorphisms (SNPs) and evaluation for genetic mapping in European pear and interspecific Pyrus hybrids

We have used new generation sequencing (NGS) technologies to identify single nucleotide polymorphism (SNP) markers from three European pear (Pyrus communis L.) cultivars and subsequently developed a subset of 1096 pear SNPs into high throughput markers by combining them with the set of 7692 apple SNPs on the IRSC apple Infinium® II 8K array. We then evaluated this apple and pear Infinium® II 9K SNP array for large-scale genotyping in pear across several species, using both pear and apple SNPs. The segregating populations employed for array validation included a segregating population of European pear (‘Old Home’בLouise Bon Jersey’) and four interspecific breeding families derived from Asian (P. pyrifolia Nakai and P. bretschneideri Rehd.) and European pear pedigrees. In total, we mapped 857 polymorphic pear markers to construct the first SNP-based genetic maps for pear, comprising 78% of the total pear SNPs included in the array. In addition, 1031 SNP markers derived from apple (13% of the total apple SNPs included in the array) were polymorphic and were mapped in one or more of the pear populations. These results are the first to demonstrate SNP transferability across the genera Malus and Pyrus. Our construction of high density SNP-based and gene-based genetic maps in pear represents an important step towards the identification of chromosomal regions associated with a range of horticultural characters, such as pest and disease resistance, orchard yield and fruit quality.

Identification of multiple key genes involved in pathogen defense and multi-stress tolerance using microarray and network analysis

This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Brassinosteroid (BR), a plant steroid hormone, plays key roles in numerous growth and developmental processes as well as tolerance to both abiotic and biotic stress. To understand the biological networks involved in BR-mediated signaling pathways and stress tolerance, we performed comparative genome-wide transcriptome analysis of a constitutively activated BR bes1-D mutant with an Agilent Arabidopsis 4 x 44K oligo chip. As a result, we newly identified 1,091 (562 up-regulated and 529 downregulated) significant differentially expressed genes (DEGs). The combination of GO enrichment and protein network analysis revealed that stress-related processes, such as metabolism, development, abiotic/biotic stress, immunity, and defense, were critically linked to BR signaling pathways. Among the identified gene sets, we confirmed more than a 6-fold up-regulation of NB-ARC and FLS2 in bes1-D plants. However, some genes, including TIR1, TSA1 and OCP3, were down-regulated. Consistently, BR-activated plants showed higher tolerance to drought stress and pathogen infection compared to wild-type controls. In this study, we newly developed a useful, comprehensive method for large-scale identification of critical network and gene sets with global transcriptome analysis using a microarray. This study also showed that gain of function in the bes1-D gene can regulate the adaptive response of plants to various stressful conditions.

Draft Genome Sequence of the Asian Pear Scab Pathogen, Venturia nashicola

Venturia nashicola, which causes scab disease of Asian pear, is a host-specific, biotrophic fungus, with a sexual stage that occurs during saprobic growth. V. nashicola is endemic to Asia and is regarded as a quarantine threat to Asian pear production outside of this continent. Currently, fungicide applications are routinely used to control scab disease. However, fungicide resistance in V. nashicola, as in other fungal pathogens, is an ongoing challenge and alternative control or prevention measures that include, for example, the deployment of durable host resistance, are required. A close relative of V. nashicola, V. pirina, causes scab disease of European pear. European pear displays non-host resistance (NHR) to V. nashicola and Asian pears are non-hosts of V. pirina. It is anticipated that the host specificity of these two fungi is governed by differences in their effector arsenals, with a subset responsible for activating NHR. The Pyrus-Venturia pathosystems provide a unique opportunity to dissect the underlying genetics of non-host interactions and to understand coevolution in relation to this potentially more durable form of resistance. Here, we present the first V. nashicola draft whole genome sequence (WGS), which is made up of 40,800 scaffolds (totalling 45 Mb) and 11,094 predicted genes. Of these genes, 1,232 are predicted to encode a secreted protein by SignalP, with 273 of these predicted to be effectors by EffectorP. The V. nashicola WGS will enable comparison to the WGSs of other Venturia spp. to identify effectors that potentially activate NHR in the pear scab pathosystems.

Lactic Acid Bacteria Isolated from Kimchi to Evaluate Anti-obesity Effect in High Fat Diet-induced Obese Mice

Obesity is one of the major public concerns due to its various side-effects such as metabolic and chronic ailments. In this study, two lactic acid bacteria (LAB) strains, Leuconostoc mesenteroides and Lactobacillus sakei, isolated from Kimchi were investigated for their anti-obesity activity in obese mice model. To induce obesity in model mice (C57BL/6), high fat diet (HFD) with 60 kcal% fat was fed for 10 weeks. LAB supplementation reduced blood urea nitrogen, glucose, and triglyceride levels in serum from obesity induced mice. The study also observed significant decrease of fibrosis, triglyceride, and total cholesterol level in fatty liver tissue by LAB supplementation. The results suggest that supplementation of L. mesenteroides and L. sakei could be used as alternative nutritional interventions to decrease symptoms caused by obesity with no safety or intolerance related problem.

Aptamer-Based Pathogen Monitoring for Salmonella enterica ser. Typhimurium

Salmonella enterica ser. Typhimurium is a foodborne pathogen that causes salmonellosis. Symptoms of salmonellosis include fever, diarrhea, and gastroenteritis. Conventional culture methods for detecting foodborne bacterial pathogens require long incubation time, expensive immunoassay methods, and sample enrichment steps. The objective of this study was to develop an aptamer-based sandwich assay to detect S. enterica ser. Typhimurium. First, S. enterica ser. Typhimurium specific binding aptamers (S11 and S24) were obtained by whole-cell SELEX for high sensitivity and specificity detection using live S. enterica ser. Typhimurium. S11 and S24 aptamers were able to capture S. enterica ser. Typhimurium selectively and distinguish it from other species of Salmonella (S. enterica ser. Typhimurium, S. enterica ser. Choleraesuis, S. enterica ser. Dublin, and S. enterica ser. Enteritidis) and food-borne bacterial pathogens (Escherichia coli K12, Listeria monocytogenes, Shigella sonnei and Staphylococcus aureus) with KD values of 4.41×10-12 M and 3.75×10-11 M, respectively. S. enterica ser. Typhimurium. aptamer-based sandwich assay exhibited a linear response for sensing S. enterica ser. Typhimurium. cells at concentration ranging from 2×101 to 2×105 CFU/mL. This aptamer-based sandwich assay can be used for on-site detection of S. enterica ser. Typhimurium.

Anti-inflammatory effect of lactic acid bacteria isolated from kimchi on acid-induced acute colitis in model mice

Lactic acid bacteria (LAB) provide numerous beneficial effects on the host body, especially on the intestine. Two LAB strains isolated from Kimchi, Leuconostoc mesenteroides and Lactobacillus sakei, were studied for its anti-inflammatory activity in acidinduced acute colitis in mice. To induce acute colitis in model mice (C57BL/6), 3% of dextran sulfate sodium treatment was treated for 7 days. Assessment of necropsy and histopathology analysis showed that oral supplementation of both L. mesenteroides and L. sakei ameliorated the symptoms of acute colitis. Moreover, the mixture of L. meseneroides and L. sakei showed synergistic effect on colitis. The results suggest that the formulation of L. mesenteroides and L. sakei mixture could be used as an oral supplementation to decrease the inflammatory harmful environment associated with colitis.

Detection of endotoxins using nanomaterials

An endotoxin is a lipopolysaccharide complex that is associated with the outer membrane of gram negative bacteria, and even a small amount can trigger strong exothermic activity. Pharmaceutical and food products can be contaminated with endotoxins, so there is a strict need to control them. A limulus amoebocyte lysate (LAL) assay is a standard endotoxin detection method that is highly sensitive and easy to use, but it has disadvantages in that the sample preparation is a complex process, it takes a long time to confirm the presence of endotoxins, and it requires a controlled workplace. Recently, endotoxin detection using nanomaterials has been gaining attention, and this review reports on the latest endotoxin detection methods using gold nanoparticles, magnetic nanoparticles, quantum dots, and carbon materials, among different nanomaterials, and it also introduces the future prospect of endotoxin detection.

Characterization of the Rosellinia necatrix Transcriptome and Genes Related to Pathogenesis by Single-Molecule mRNA Sequencing

White root rot disease, caused by the pathogen Rosellinia necatrix, is one of the world’s most devastating plant fungal diseases and affects several commercially important species of fruit trees and crops. Recent global outbreaks of R. necatrix and advances in molecular techniques have both increased interest in this pathogen. However, the lack of information regarding the genomic structure and transcriptome of R. necatrix has been a barrier to the progress of functional genomic research and the control of this harmful pathogen. Here, we identified 10,616 novel full-length transcripts from the filamentous hyphal tissue of R. necatrix (KACC 40445 strain) using PacBio single-molecule sequencing technology. After annotation of the unigene sets, we selected 14 cell cycle-related genes, which are likely either positively or negatively involved in hyphal growth by cell cycle control. The expression of the selected genes was further compared between two strains that displayed different growth rates on nutritional media. Furthermore, we predicted pathogen-related effector genes and cell wall-degrading enzymes from the annotated gene sets. These results provide the most comprehensive transcriptomal resources for R. necatrix, and could facilitate functional genomics and further analyses of this important phytopathogen.

Breeding of ‘Joyskin’ Pear as fruit for Eating withthe Skin

In 1994, a new cultivar ‘Joyskin’ was created from a cross between the cultivars ‘Whangkeumbae’ and ‘Waseaka’ at the Pear Research Institute of the National Institute of Horticultural and Herbal Science, Rural Development Administration. In 2006, the ‘Joyskin’ was selected from among the 317 seedlings resulting from the cross for its skin and taste qualities. Regional adaptation tests were conducted in nine regions and in ten experimental plots from 2006 to 2011. The cultivar was named in 2011. ‘Joyskin’ showed a vigorous growth habit and semi-spread characteristics similar to ‘Whangkeumbae’. The average full bloom date for ‘Joyskin’ was April 21st, which was also similar to ‘Whangkeumbae’. The optimum fruit ripening time was September 6-8th, which was six or eight days earlier than ‘Whangkeumbae’. The fruit was round in shape and the skin was a golden yellow color at maturity. The average fruit weight was 320 g and the flesh firmness was 2.5 kg/8mmφ. The firmness of the fruit skin determined by a blade-type plunger of texture analyzer was 22.9 N, which was significantly different from that of ‘Whangkeumbae’ 29.9N. Stone cell analysis of ‘Joyskin’ by phloroglucinol-HCl, showed that ‘Joyskin’ stone cells were small in size and few in numbers cpmpared to those of cultivars of was ‘Manpungbae’, ‘Niitaka’, and ‘Whangkeumbae’. The patent application for ‘Joyskin’ was submitted in April, 2012 (Grant No. 2012-337). In 2016, ‘Joyskin’ (Grant No. 5895) was registered as a separate record, with uniformity and stability per Korean Seed Industry Law. Additional key words: crossing breeding, fruit, Pyrus pyrifolia, self-incompatibility, stone cell