• Journal of Microbiology and Biotechnology
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• v.24 no.6
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• pp.748-755
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• 2014

In the genome annotation of Escherichia coli MG1655, the orf382 (1,149 bp) is designated as a gene encoding an alcohol dehydrogenase that may be Fe-dependent. In this study, the gene was amplified from the genome by PCR and overexpressed in Escherichia coli BL21(DE3). The recombinant $6{\times}$His-tag protein was then purified and characterized. In an enzymatic assay using different hydroxyl-containing substrates (n-butanol, $\small{L}$-threonine, ethanol, isopropanol, glucose, glycerol, $\small{L}$-serine, lactic acid, citric acid, methanol, or $\small{D}$-threonine), the enzyme showed the highest activity on $\small{L}$-threonine. Characterization of the mutant constructed using gene knockout of the orf382 also implied the function of the enzyme in the metabolism of $\small{L}$-threonine into glycine. Considering the presence of tested substrates in living E. coli cel ls and previous literature, we believed that the suitable nomenclature for the enzyme should be an $\small{L}$-threonine dehydrogenase (LTDH). When using $\small{L}$-threonine as the substrate, the enzyme exhibited the best catalytic performance at $39^{\circ}C$ and pH 9.8 with $NAD^+$ as the cofactor. The determination of the Km values towards $\small{L}$-threonine (Km = $11.29{\mu}M$), ethanol ($222.5{\mu}M$), and n-butanol ($8.02{\mu}M$) also confirmed the enzyme as an LTDH. Furthermore, the LTDH was shown to be an ion-containing protein based on inductively coupled plasma-atomic emission spectrometry with an isoelectronic point of pH 5.4. Moreover, a circular dichroism analysis revealed that the metal ion was structurally and enzymatically essential, as its deprivation remarkably changed the ${\alpha}$-helix percentage (from 12.6% to 6.3%).

• Molecules and Cells
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• v.28 no.6
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• pp.529-536
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• 2009

Genome sequencing of the pig is being accelerated because of its importance as an evolutionary and biomedical model animal as well as a major livestock animal. However, information on expressed porcine genes is insufficient to allow annotation and use of the genomic information. A series of expressed sequence tags of 5' ends of five full-length enriched cDNA libraries (SUSFLECKs) were functionally characterized. SUSFLECKs were constructed from porcine abdominal fat, induced fat cells, loin muscle, liver, and pituitary gland, and were composed of non-normalized and normalized libraries. A total of 55,658 ESTs that were sequenced once from the 5′ ends of clones were produced and assembled into 17,684 unique sequences with 7,736 contigs and 9,948 singletons. In Gene Ontology analysis, two significant biological process leaf nodes were found: gluconeogenesis and translation elongation. In functional domain analysis based on the Pfam database, the beta transducin repeat domain of WD40 protein was the most frequently occurring domain. Twelve genes, including SLC25A6, EEF1G, EEF1A1, COX1, ACTA1, SLA, and ANXA2, were significantly more abundant in fat tissues than in loin muscle, liver, and pituitary gland in the SUSFLECKs. These characteristics of SUSFLECKs determined by EST analysis can provide important insight to discover the functional pathways in gene networks and to expand our understanding of energy metabolism in the pig.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.3
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• pp.320-333
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• 2019

Objective: The Shanghai Holstein cattle breed is susceptible to severe mastitis and other diseases due to the hot weather and long-term humidity in Shanghai, which is the main distribution centre for providing Holstein semen to various farms throughout China. Our objective was to determine the genetic mechanisms influencing economically important traits, especially diseases that have huge impact on the yield and quality of milk as well as reproduction. Methods: In our study, we detected the structural variations of 1,092 Shanghai Holstein cows by using next-generation sequencing. We used the DELLY software to identify deletions and insertions, cn.MOPS to identify copy-number variants (CNVs). Furthermore, we annotated these structural variations using different bioinformatics tools, such as gene ontology, cattle quantitative trait locus (QTL) database and ingenuity pathway analysis (IPA). Results: The average number of high-quality reads was 3,046,279. After filtering, a total of 16,831 deletions, 12,735 insertions and 490 CNVs were identified. The annotation results showed that these mapped genes were significantly enriched for specific biological functions, such as disease and reproduction. In addition, the enrichment results based on the cattle QTL database showed that the number of variants related to milk and reproduction was higher than the number of variants related to other traits. IPA core analysis found that the structural variations were related to reproduction, lipid metabolism, and inflammation. According to the functional analysis, structural variations were important factors affecting the variation of different traits in Shanghai Holstein cattle. Our results provide meaningful information about structural variations, which may be useful in future assessments of the associations between variations and important phenotypes in Shanghai Holstein cattle. Conclusion: Structural variations identified in this study were extremely different from those of previous studies. Many structural variations were found to be associated with mastitis and reproductive system diseases; these results are in accordance with the characteristics of the environment that Shanghai Holstein cattle experience.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2019.10a
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• pp.73-73
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• 2019

Angelica gigas Nakai (Korean danggui), a member of the Umbelliferae family, is a Korean traditional medicinal plant whose roots have been used for treating gynecological diseases. Transcriptomics is the study of the transcriptome, which is the complete set of RNA transcripts that are produced by the genome, using high-throughput methods, such as microarray analysis. In this study, transcriptome analysis of A.gigas Nakai was carried out. Transcriptome sequencing and assembly was carried out by using Illumina Hiseq 2500, Velvet and Oases. A total of 109,591,555 clean reads of A. gigas Nakai was obtained after trimming adaptors. The obtained reads were assembled with an average length of 1,154 bp, a maximum length of 13,166 bp, a minimum length of 200 pb, and N50 of 1,635 bp. Functional annotation and classification was performed using NCBI NR, InterprotScan, KOG, KEGG and GO. Candidate genes for phenylpropanoid biosynthesis were obtanied from A.gigas transcriptome and the genes and its proteins were confirmed through the NCBI homology BLAST searches, revealing high identity with other othologous genes and proteins from various plants pecies. In RNA sequencing analysis using an Illumina Next-Seq2500 sequencer, we identified a total 94,930 transcripts and annotated 71,281 transcripts, which provide basic information for further research in A.gigas Nakai. Our transcriptome data reveal that several differentially expressed genes related to flower color in A.gigas Nakai. The results of this research provide comprehensive information on the A.gigas Nakai genome and enhance our understanding of the flower color related gene pathways in this plant.

• Plant Breeding and Biotechnology
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• v.5 no.4
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• pp.334-343
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• 2017

Eclipta prostrata and E. alba are annual herbal medicinal plants and have been used as Chinese medicinal tonics. Both species are widely distributed in tropical and subtropical regions as well as in Korea. Both species have similar morphological features but E. alba has smoother leaf blade margins compared with E. prostrata. Although both species are utilized as oriental medicines, E. prostrata is more widely used than E. alba. Morphological semblances have confounded identification of either species. Here, we report the complete chloroplast genomes of both species to provide an authentication system between the two species and understand their diversity. Both chloroplast genomes were 151,733-151,757 bp long and composed of a large single copy (83,285-83,300 bp), a small single copy (18,283-18,346 bp), and a pair of inverted repeats (25,075-25,063 bp). Gene annotation revealed 80 protein coding genes, 30 tRNA genes and four rRNA genes. A phylogenetic analysis revealed that the genus Eclipta is grouped with Heliantheae tribe species in the Asteraceae family. A comparative analysis verified 29 InDels and 58 SNPs between chloroplast genomes of E. prostrata and E. alba. The low chloroplast genome sequence diversity indicates that both species are really close to each other and are not completely diverged yet. We developed six DNA markers that distinguish E. prostrata and E. alba based on the polymorphisms of chloroplast genomes between E. prostrata and E. alba. The chloroplast genome sequences and the molecular markers generated in this study will be useful for further research of Eclipta species and accurate classification of medicinal herbs.

• Molecular & Cellular Toxicology
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• v.3 no.3
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• pp.208-213
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• 2007

With the help of a development and popularization of microarray technology that enable to us to simultaneously investigate the expression pattern of thousands of genes, the toxicogenomics experimenters can interpret the genome-scale interaction between genes exposed in toxicant or toxicant-related environment. The ultimate and primary goal of toxicogenomics identifies functional context among the group of genes that are differentially or similarly coexpressed under the specific toxic substance. On the other side, public reference databases with transcriptom, proteom, and biological pathway information are needed for the analysis of these complex omics data. However, due to the heterogeneous and independent nature of these databases, it is hard to individually analyze a large omics annotations and their pathway information. Fortunately, several web sites of the public database provide information linked to other. Nevertheless it involves not only approriate information but also unnecessary information to users. Therefore, the systematically integrated database that is suitable to a demand of experimenters is needed. For these reasons, we propose SOP (Search of Omics Pathway) database system which is constructed as the integrated biological database converting heterogeneous feature of public databases into combined feature. In addition, SOP offers user-friendly web interfaces which enable users to submit gene queries for biological interpretation of gene lists derived from omics experiments. Outputs of SOP web interface are supported as the omics annotation table and the visualized pathway maps of KEGG PATHWAY database. We believe that SOP will appear as a helpful tool to perform biological interpretation of genes or proteins traced to omics experiments, lead to new discoveries from their pathway analysis, and design new hypothesis for a next toxicogenomics experiments.

• ALGAE
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• v.35 no.2
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• pp.177-187
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• 2020

Cyanobacteria have been widely reported to produce a variety of UV-absorbing mycosporine-like amino acids (MAAs). Herein, we reported production of the unusual MAA, mycosporine-glycine-alanine (MGA) in the cyanobacterium Sphaerospermopsis torques-reginae ITEP-024 using a newly developed UHPLC-DAD-MS/HRMS (ultra-high performance liquid chromatography-diode array detection-high resolution tandem mass spectrometry) method. MGA had previously been first identified in a red-algae, but S. torques-reginae strain ITEP-024 is the first cyanobacteria to be reported as an MGA producer. Herein, the chemical structure of MGA is fully elucidated from one-dimensional / two-dimensional nuclear magnetic resonance and HRMS data analyses. MAAs are unusually produced constitutively in S. torques-reginae ITEP-024, and this production was further enhanced following UV-irradiance. It has been proposed that MAA biosynthesis proceeds in cyanobacteria from the pentose phosphate pathway intermediate sedoheptulose 7-phosphate. Annotation of a gene cluster encoded in the genome sequence of S. torques-reginae ITEP-024 supports these gene products could catalyse the biosynthesis of MAAs. However, addition of glyphosate to cultures of S. torques-reginae ITEP-024 abolished constitutive and ultra-violet radiation induced production of MGA, shinorine and porphyra-334. This finding supports involvement of the shikimic acid pathway in the biosynthesis of MAAs by this species.

• Food Science of Animal Resources
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• v.35 no.5
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• pp.692-702
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• 2015

Lactobacillus mucosae is a natural resident of the gastrointestinal tract of humans and animals and a potential probiotic bacterium. To understand the global protein expression profile and metabolic features of L. mucosae LM1 in the early stationary phase, the QExactive^TM Hybrid Quadrupole-Orbitrap Mass Spectrometer was used. Characterization of the intracellular proteome identified 842 proteins, accounting for approximately 35% of the 2,404 protein-coding sequences in the complete genome of L. mucosae LM1. Proteome quantification using QExactive^TM Orbitrap MS detected 19 highly abundant proteins (> 1.0% of the intracellular proteome), including CysK (cysteine synthase, 5.41%) and EF-Tu (elongation factor Tu, 4.91%), which are involved in cell survival against environmental stresses. Metabolic pathway annotation of LM1 proteome using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database showed that half of the proteins expressed are important for basic metabolic and biosynthetic processes, and the other half might be structurally important or involved in basic cellular processes. In addition, glycogen biosynthesis was activated in the early stationary phase, which is important for energy storage and maintenance. The proteogenomic data presented in this study provide a suitable reference to understand the protein expression pattern of lactobacilli in standard conditions

• Food Science of Animal Resources
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• v.40 no.5
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• pp.746-757
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• 2020

Enterotoxigenic Escherichia coli (ETEC) is the major pathogenic E. coli that causes diarrhea and edema in post-weaning piglets. In this study, we describe the morphology and characteristics of ØCJ19, a bacteriophage that infects ETEC, and performed genetic analysis. Phage ØCJ19 belongs to the family Myoviridae. One-step growth curve showed a latent phase of 5 min and burst size of approximately 20 phage particles/infected cell. Phage infectivity was stable for 2 h between 4℃ and 55℃, and the phage was stable between pH 3 and 11. Genetic analysis revealed that phage ØCJ19 has a total of 49,567 bases and 79 open reading frames (ORFs). The full genomic sequence of phage ØCJ19 showed the most similarity to an Escherichia phage, vB_EcoS_ESCO41. There were no genes encoding lysogeny, toxins, virulence factors, or antibiotic resistance in this phage, suggesting that this phage can be used safely as a biological agent to control ETEC. Comparative genomic analysis in terms of the tail fiber proteins could provide genetic insight into host recognition and the relationship with other coliphages. These results showed the possibility to improve food safety by applying phage ØCJ19 to foods of animal origin contaminated with ETEC and suggests that it could be the basis for establishing a safety management system in the animal husbandry.

• Journal of Plant Biotechnology
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• v.41 no.1
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• pp.26-32
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• 2014

Recent genome annotation revealed that Populus trichocarpa contains 81 glutathione S-transferase (GST) genes. GST genes play important and varying roles in plants, including conferring tolerance to various abiotic stresses. Little information is available on the relationship - if any - between drought/salt stresses and GSTs in woody plants. In this study, we screened the PatgGST genes in hybrid poplar (Populus alba ${\times}$ Populus tremula var. glandulosa) that were predicted to confer drought tolerance based on our expression analysis of all members of the poplar GST superfamily following exposure to salt (NaCl) and drought (PEG) stresses, respectively. Exposure to the salt stress resulted in the induction of eight PatgGST genes and down-regulation of one PatgGST gene, and the level of induction/repression was different in leaf and stem tissues. In contrast, 16 PatgGST genes were induced following exposure to the drought (PEG) stress, and two were down-regulated. Taken together, we identified seven PatgGSTs (PatgGSTU15, PatgGSTU18, PatgGSTU22, PatgGSTU27, PatgGSTU46, PatgGSTU51 and PatgGSTU52) as putative drought tolerance genes based on their induction by both salt and drought stresses.