• Proceedings of the Korean Society of Life Science Conference
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• 2001.06a
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• pp.67-86
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• 2001

A strain producing strongly fibrinolytic enzyme was isolated from soil and was identified to be Bacillus subtilis by biochemical and physiological characterization. The optimal culture conditions for the production of fibrinolytic enzyme was determined to be 1.0% tryptone, 1.5% soluble starch, 0.5% Peptone, 0.5% NaCl, $(NH_{4})_{3}PO_4.3H_{2}O, and MgSO_{4}.7H_{2}O.$ Initial pH and temperature were pH 8.0 and $30^{\circ}C$ , respectively, The highest enzyme production was observed at 30 hours of cultivation at $30^{\circ}C$ The fibrinolytic enzyme was purified to homogeneity by DEAE Sephadex A-50 ion exchange column chromatography, 70% ammonium sulfate precipitation, Sephadex G-200 and G-75 gel filtration column chromatography. The molecular weight of the purified enzyme was 28,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A gene encoding the fibrinolytic enzyme was cloned into a plasmid vector pBluescript, transforming E.coli XL-1 Blue. The clone was able to degrade fibrin, This indicated that the gene could encode a fibrinolytic enzyme. The nucleotide sequence of the 2.7 kb insert was determined in both direction. One open reading frame composed of 1023 nucleotides was found to be a potential protein coding region. There was the putative Shine-Dalgano sequence and TATA box upstream of the open reading frame. The homology search data in the genome database showed that both the 2.7 kb insert and 1 kb open reading frame carried no significance in the nucleotide sequence of known fibrinolytic enzyme from Bacillus serovars. The recombinant cell harboring the novel gene involved in fibrinolysis was subjected to protein purification. The molecular mass of the purified fibrinolytic enzyme was determined to be 31864 Dalton, which was highly in accordance with the molecular mass(33 kDa) of the fibrinolytic gene deduced from the insert. The fibrinolytic enzyme was Purified 50.5 folds to homogeneity in overall yield of 10.7% by DEAE Sephadex A-50 ion exchange, 85% ammonium sulfate precipitation, Sephadex G-50, Superdex 75 HR FPLC gel filtration. In conclusion, a novel fibrinolytic gene from Bacillus subtilis was identified and characterized by cloning a genomic library of Bacillus subtilis into pBleuscript. For the soybean fermented by this strain, it is found that there increased assistant protein about 20% compared to the soybean not fermented and increased about 30% according to amino acid analysis and, in particular, essential amino acid increased about 40%. When keeping this fermented soybean powder at room temperature for about 70days, it showed very high stability maintaining almost perfect activity and, therefore, it gave us great suggestion its possibility of development as a new functional food.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.3
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• pp.279-286
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• 2005

Objective: Although several genetic factors have been associated with defects in human spermatogenesis, the unambiguous causative genes have not been elucidated. The male infertility by haploinsufficiency of PRM1 or PRM2 has been reported in mouse model. The aim of this study was to identify the single nucleotide polymorphisms (SNPs) of PRM1 and PRM2, related to the genotype of Korean infertile men. Methods: Genomic DNAs were extracted from peripheral bloods of infertile men with oligozoospermia or azoospermia, and analyzed using polymerase chain reaction (PCR) and direct sequencing. We carried out the direct sequencing analysis of amplified fragments in PRM1 (557 nucleotides from -42 to 515) and PRM2 (599 nucleotides from 49 to 648) genes, respectively. Results: Three SNPs of coding region in the PRM1 gene was found in the analysis of 130 infertile men. However, the SNPs at a133g (aa 96.9%, ag 3.1% and gg 0.0%), c160a (cc 99.2%, ca 0.8% and aa 0.0%) and c321a (cc 56.9%, ca 35.4% and aa 7.7%) coded the same amino acids, in terms of silence phenotypes. On the other hand, as results of the PRM2 gene sequencing in 164 infertile men, only two SNPs, g398c (gg 62.2%, gc 31.1% and ga 6.7%) and a473c (aa 63.4%, ac 29.9% and cc 6.7%), were identified in the intron of the PRM2 gene. Conclusions: There was no mutation and significant SNPs on PRM1 and PRM2 gene in Korean infertile men. These results suggest that the PRM1 and PRM2 genes are highly conserved and essential for normal fertility of men.

• The Korean Journal of Malacology
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• v.25 no.3
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• pp.223-230
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• 2009

Metallothioneins (MTs) play a key role in metallic homeostasis and detoxification in most living organisms. In an attempt to study the biological functions and significance of MT in a snail, we cloned and partially characterized the MT gene from the left-handed snail, Physa acutawhich has been regarded as a potential biomonitering species for fresh water. The complete cDNA sequence of PaMT cDNA was identified from the expressed sequence tag (EST) sequencing project of Physa acuta. The coding region of 180 bp gives 60 amino acid residues including the initiation methionine and termination codon. Clustering and phylogenic analysis of PaMT with other MT amino acid sequences show that it has some identities to Helix pomatia (60%), Arianta arbustorum (58%), Perna viridis (49%), Mytilus edulis (49%), Bathymodiolus azoricus (49%), Bathymodiolus azoricus (48%) and Bathymodiolus sp. FD-2002 (48%). Time dependent induction for PaMT from P. acuta exposed with cadmium (50 ppb) indicated that PaMT was induced at 4-8 hr after exposure. It remains to further develop PaMTas a potential biomarker for water contamination in fresh water.

• The Korean Journal of Malacology
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• v.29 no.3
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• pp.171-179
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• 2013

Arginine kinase (ArK) is known to play an important role in most invertebrates the level of ATP by phosphorylation of phosphagens in cell and immuninty in living organisms. ArK has been identified in many kinds of organisms ranging from invertebrate to vertebrate. However, no ArK gene has been cloned and investigated from N. samarangae. This leads us to identify ArK cDNA (NsArK) from the expressed sequence tag (EST) sequencing of N. samarangae. Sequence analysis indicated that the coding region of 1,065 bp contains 355 amino acid residues. Molecular phylogenetic analysis shows that NsArK had very high similarities with mollusca and arthropoda. In an attempt to investigate a potential role of NsArK in the digestive gland of N. samarangae, expression patterns were analyzed. RT-PCR analsysis shows that NsArK mRNA is induced in the rane of 1.2 fold at 6 hr by laminarin when compared with the control. The immunnologial and physiological role of NsArK remains to be further investigated in N. samarangae.

• Korean Journal of Agricultural Science
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• v.46 no.3
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• pp.507-517
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• 2019

To follow up on a 2017 survey of tomato virus diseases, samples with virus-like symptoms were collected from the same areas (Buyeo-gun, Chungchungnam-Do and Daejeon, Korea) in 2018. While in 2017 mixed infections of Tomato mosaic virus with either Tomato yellow leaf curl virus (TYLCV) or Tomato chlorosis virus were detected, only TYLCV was detected in symptomatic samples in 2018. TYLCV amplicons of c.777 bp representing the coat protein (CP) coding region were cloned from the TYLCV positive samples, and the sequence data showed a 97.17% to 98.84% nucleotide and 98.45% to 99.22% amino acid identity with the 2017 Buyeo-gun isolate (MG787542), which had the highest amino acid (aa) sequence identity of up to 99.2% with four 2018 Buyeo-gun sequences (MK521830, MK521833, MK521834, and MK521835). The lowest aa sequence identity of 98.45% was found in a 2018 Daejeon isolate (MK521836); the distance between Buyeo-gun and Daejeon is about 45 km. Phylogenetic analysis indicated that the currently reported CP sequences are most closely related to Korean sequences from Masan (HM130912), Goseong (JN680149), Busan (GQ141873), Boseong (GU325634), and the 2017 isolate TYLCV-N (MG787543) in the 'Japan' cluster of TYLCV isolates and distinct from the 'China' cluster isolates from Nonsan (GU325632), Jeonju (HM130913) and Jeju (GU325633, HM130914). Our survey data from 2017 and 2018 suggest that TYLCV has become established in Korea and may be spread by whitefly vectors from weed reservoirs within the farm environment.

• Journal of Animal Science and Technology
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• v.60 no.12
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• pp.32.1-32.10
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• 2018

Background: Necdin (NDN), a member of the melanoma antigen family showing imprinted pattern of expression, has been implicated as causing Prader-Willi symptoms, and known to participate in cellular growth, cellular migration and differentiation. The region where NDN is located has been associated to QTLs affecting reproduction and early growth in cattle, but location and functional analysis of the molecular mechanisms have not been established. Methods: Here we report the sequence variation of the entire coding sequence from 72 samples of cattle, yak, buffalo, goat and sheep, and discuss its variation in Bovidae. Median-joining network analysis was used to analyze the variation found in the species. Synonymous and non-synonymous substitution rates were determined for the analysis of all the polymorphic sites. Phylogenetic analysis were carried out among the species of Bovidae to reconstruct their relationships. Results: From the phylogenetic analysis with the consensus sequences of the studied Bovidae species, we found that only 11 of the 26 nucleotide changes that differentiate them produced amino acid changes. All the SNPs found in the cattle breeds were novel and showed similar percentages of nucleotides with non-synonymous substitutions at the N-terminal, MHD and C-terminal (12.3, 12.8 and 12.5%, respectively), and were much higher than the percentage of synonymous substitutions (2.5, 2.6 and 4.9%, respectively). Three mutations in cattle and one in sheep, detected in heterozygous individuals were predicted to be deleterious. Additionally, the analysis of the biochemical characteristics in the most common form of the proteins in each species show very little difference in molecular weight, pI, net charge, instability index, aliphatic index and GRAVY (Table 4) in the Bovidae species, except for sheep, which had a higher molecular weight, instability index and GRAVY. Conclusions: There is sufficient variation in this gene within and among the studied species, and because NDN carry key functions in the organism, it can have effects in economically important traits in the production of these species. NDN sequence is phylogenetically informative in this group, thus we propose this gene as a phylogenetic marker to study the evolution and conservation in Bovidae.

• Convergence Security Journal
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• v.19 no.2
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• pp.91-104
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• 2019

In this paper, we present a novel local descriptor, Local Prominent Directional Pattern (LPDP), to represent the description of facial images for gender recognition purpose. To achieve a clearly discriminative representation of local shape, presented method encodes a target pixel with the prominent directional variations in local structure from an analysis of statistics encompassed in the histogram of such directional variations. Use of the statistical information comes from the observation that a local neighboring region, having an edge going through it, demonstrate similar gradient directions, and hence, the prominent accumulations, accumulated from such gradient directions provide a solid base to represent the shape of that local structure. Unlike the sole use of gradient direction of a target pixel in existing methods, our coding scheme selects prominent edge directions accumulated from more samples (e.g., surrounding neighboring pixels), which, in turn, minimizes the effect of noise by suppressing the noisy accumulations of single or fewer samples. In this way, the presented encoding strategy provides the more discriminative shape of local structures while ensuring robustness to subtle changes such as local noise. We conduct extensive experiments on gender recognition datasets containing a wide range of challenges such as illumination, expression, age, and pose variations as well as sketch images, and observe the better performance of LPDP descriptor against existing local descriptors.

• The Plant Pathology Journal
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• v.38 no.5
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• pp.541-549
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• 2022

Potato late blight caused by Phytophthora infestans is a destructive disease in Korea. To elucidate the genomic variation of the mitochondrial (mt) genome, we assembled its complete mt genome and compared its sequence among different haplotypes. The mt genome sequences of four Korean P. infestans isolates were revealed by Illumina HiSeq. The size of the circular mt genome of the four major genotypes, KR_1_A1, KR_2_A2, SIB-1, and US-11, was 39,872, 39,836, 39,872, and 39,840 bp, respectively. All genotypes contained the same 61 genes in the same order, comprising two RNA-encoding genes, 16 ribosomal genes, 25 transfer RNA, 17 genes encoding electron transport and ATP synthesis, 11 open reading frames of unknown function, and one protein import-related gene, tatC. The coding region comprised 91% of the genome, and GC content was 22.3%. The haplotypes were further analyzed based on sequence polymorphism at two hypervariable regions (HVRi), carrying a 2 kb insertion/deletion sequence, and HVRii, carrying 36 bp variable number tandem repeats (VNTRs). All four genotypes carried the 2 kb insertion/deletion sequence in HVRi, whereas HVRii had two VNTRs in KR_1_A1 and SIB-1 but three VNTRs in US-11 and KR_2_A2. Minimal spanning network and phylogenetic analysis based on 5,814 bp of mtDNA sequences from five loci, KR_1_A1 and SIB-1 were classified as IIa-6 haplotype, and isolates KR_1_A2 and US-11 as haplotypes IIa-5 and IIb-2, respectively. mtDNA sequences of KR_1_A1 and SIB-1 shared 100% sequence identity, and both were 99.9% similar to those of KR_2_A2 and US-11.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.84-84
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• 2003

DNA methylation is involved in epigenetic processes such as X-chromosome inactivation, imprinting and silencing of transposons. DNA methylation is a highly plastic and critical component of mammalian development The DNA methyltransferases (Dnmts) are responsible for the generation of genomic methylation patterns, which lead to transcriptional silencing. The maintenance DNA methyltransferase enzyme, Dnmt 1, and the de novo methyltransferase, Dnmt3a and Dnmt3b, are indispensable for development because mice homozygous for the targeted disruption of any of these genes are not viable. The occurrence of DNA methylation is not random, and it can result in gene silencing The mechanisms underlying these processes are poorly understood. It is well established that DNA methylation and histone deacetylation operate along a common mechanistic pathway to repress transcription through the action of methyl-binding domain proteins (MBDs), which are components of, or recruit, histone deacetylase (HDAC) complexes to methylated DNA. As a basis for future studies on the role of the DNA-methyl-transferase in porcine development, we have isolated and characterized a partial cDNA coding for the porcine Dnmt1. Total RNA of testis, lung and ovary was isolated with TRlzol according to the manufacture's specifications. 5 ug of total RNA was reverse transcribed with Super Script II in the presence of porcine Dnmt 1 specific primers. Standard PCRs were performed in a total volume of 50 ul with cDNA as template. Two DNA fragmenets in different position were produced about 700bp, 1500bp and were cloned into pCR II-TOPO according to the manufacture's specification. Assembly of all sequences resulted in a cDNA from 158bp of 5'to 4861bp of 3'compare with the known human maintenance methyltransferase. Now, we are cloning the unknown Dnmt 1 region by 5'-RACE method and expression of Dnmt 1 in tissues from adult porcine animals.

• Animal Bioscience
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• v.37 no.7
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• pp.1156-1167
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• 2024

Objective: MicroRNAs (miRNAs) are endogenous non-coding RNAs that can play a role in the post-transcriptional regulation of mammalian preadipocyte differentiation. However, the precise functional mechanism of its regulation of fat metabolism is not fully understood. Methods: We identified bta-miR-365-3p, which specifically targets the 3' untranslated region (3'UTR) of the FK506-binding protein 5 (FKBP5), and verified its mechanisms for regulating expression and involvement in adipogenesis. Results: In this study, we found that the overexpression of bta-miR-365-3p significantly decreased the lipid accumulation and triglyceride content in the adipocytes. Compared to inhibiting bta-miR-36 5-3p group, overexpression of bta-miR-365-3p can inhibit the expression of adipocyte differentiation-related genes C/EBPα and PPARγ. The dual-luciferase reporter system further validated the targeting relationship between bta-miR-365-3p and FKBP5. FKBP5 mRNA and protein expression were detected by quantitative real-time polymerase chain reaction and Western blot. Overexpression of bta-miR-365-3p significantly down-regulated FKBP5 expression, while inhibition of bta-miR-365-3p showed the opposite, indicating that bta-miR-365-3p negatively regulates FKBP5. Adenosine 5'-monophosphate (AMP)-activated protein kinase/mammalian target of rapamycin (AMPK/mTOR) signaling pathway is closely related to the regulation of cell growth and is involved in the development of bovine adipocytes. In this study, overexpression of bta-miR-365-3p significantly inhibited mRNA and protein expression of AMPK, mTOR, and SREBP1 genes, while the inhibition of bta-miR-365-3p expression was contrary to these results. Overexpression of FKBP5 significantly upregulated AMPK, mTOR, and SREBP1 gene expression, while inhibition of FKBP5 expression was contrary to the above experimental results. Conclusion: In conclusion, these results indicate that bta-miR-365-3p may be involved in the AMPK/mTOR signaling pathway in regulating Yanbian yellow cattle preadipocytes differentiation by targeting the FKBP5 gene.