• Journal of Animal Science and Technology
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• v.48 no.5
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• pp.637-650
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• 2006

Genomic DNA isolated from two crucian carp species obtained from Yesan (Carassius auratus) and Dangjin (Carassius cuvieri) in Korea were amplified at several times by polymerase chain reaction (PCR). The amplified products were separated by agarose gel electrophoresis (AGE) with oligonucleotides decamer primer and stained with ethidium bromide. The seven arbitrarily selected primers OPC-11, OPC-14, OPC-18, OPD- 02, OPD-11, OPD-15 and OPD-20 generated the shared loci by each species, the polymorphic and specific loci. The seven primers generated the total 458 loci that can be scored from the crucian carp obtained in C. auratus species. 358 fragments were generated from the species obtained in C. cuvieri species. The size of DNA fragments varies from 150 to 1,600bp. The complexity of the banding patterns varies dramatically between the primers and two locations. In this study, 458 loci were identified in the crucian carp species from Yesan and 358 in the crucian carp species from Dangjin: 84 polymorphic loci (18.3%) in the C. auratus species and 48 (13.4%) in the C. cuvieri species. 154 shared loci by each species, the average 22 per primer, were observed in the C. auratus species and 187 loci, the average 26.7 per primer, in the Dangjin species. Based on the average bandsharing (BS) values of all samples, the similarity matrix ranged from 0.434 to 0.868 in the C. auratus species and from 0.449 to 0.924 in the C. cuvieri species. The average BS value was 0.641±0.013 within the C. auratus species and 0.684±0.013 within the C. cuvieri species. The average BS value between two crucian carp species 0.484 ± 0.007, ranged from 0.307 to 0.682. The BS value between the individual No. 09 and No. 16 was 0.682, which was the highest between two crucian carp species. Compared separately, the BS value of individuals within the C. cuvieri species was higher than the C. auratus species. The dendrogram obtained by the seven primers, indicates three genetic clusters: cluster 1 (AURATUS No. 01, 02, 03, 04, 05, 06, 07, 08, 09, 10 and 11), cluster 2 (CUVIERI No. 12, 13, 14, 15, 16, 17, 18, 19, 20 and 21) and cluster 3 (CUVIERI no. 22). The shortest genetic distance displaying significant molecular difference was between the individual AURATUS No. 09 and AURATUS No. 08 from Yesan (genetic distance=0.064). The longest genetic distance displaying significant molecular differences was between the individual CUVIERI No. 17 and AURATUS No. 11 between two crucian carp species (0.477). RAPD-PCR analysis has revealed the significant genetic distance between two crucian carp species pairs.(Key words: Carassius auratus, Carassius cuvieri, Crucian Carp, DNA Polymorphism, Genetic Distance)

• Journal of Gastric Cancer
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• v.5 no.3 s.19
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• pp.191-199
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• 2005

Background: Though infections of Helicobacter pylori (H. pylori) are closely associated with activation of host angiogenesis, the underlying mechanisms, as well as the strategy for its prevention, have not been identified. Here, we investigated a causal role of H. pylori infection in angiogenesis of gastric mucosa and a potent inhibitory effect of a gastric proton pump inhibitor (PPI) on the gastropathy. Materials and Methods: A comparative analysis of CD 34 expression in tissues obtained from 20 H. pylori-associated gastritis and 18 H. pylori-negative gastritis patients was performed. Expression of $HIF-1{\alpha}$ and VEGF were tested by using RT-PCR. To evaluate the direct effect of H. pylori infection on differentiation of endothelial HUVEC cells, we carried out an in vitro angiogenesis assay. Results: H. pyfori-associated gastritis tissues showed significantly higher density of $CD34^+$ blood vessels than did H. pylori-negative gastritis tissues, and the levels were well correlated with expressions of $HIF-1{\alpha}$. Conditioned media from H. pylori-infected gastric mucosal cells stimulated a tubular formation of HUVEC cells. We also found a significant inhibitory effect of PPI, an agent frequently used for H. pylori eradication, on H. pylori-induced angiogenesis. This drug effectively inhibited the phosphorylation of MAP kinase ERK1/2, which is a principal signal for H. pylori-induced angiogenesis. Conclusion: The fact that PPls can down-regulate H. pylori-induced angiogenesis suggest that anti-angiogenic treatment using PPI may be a preventive approach for H. pylori-associated carcinogenesis.

• Tuberculosis and Respiratory Diseases
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• v.40 no.6
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• pp.653-658
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• 1993

Background: Recent advancement of molecular genetics has revealed that malignant transformation of a cell may be a complex multistep process and this process is grouped, in general, into two distinct categories, activation of protooncogenes and inactivation of tumor suppressor genes. This study was focused on the mutation of p53 tumor suppressor gene, because p53 gene mutation is now generally accepted to be one of the most frequent genetic changes in a variety of human cancers. Although lung cancer is one of the common cancers in Korea, the genetic change in the carcinogenesis process is not yet known clearly. To investigate the role of p53 gene mutation in lung cancer, we examined the mutations of exon 4-8 of the p53 gene in humna lung cancer cell lines, because most of the mutations of p53 gene have been reported to develop in exon 4-8. Method: Genomic DNA was obtained by the digestion of proteinase K and the extraction by phenol-chloroform-ethanol method from two human pulmonary adenocarcinoma cell lines, PC-9 and PC-14, and one human small cell lung cancer cell line, H69. To detect the mutations of exon 4-8 of the p53 gene, polymerase chain reaction single-strand conformation polymorphism(PCR-SSCP) analysis was performed with the DNA extracted from the cells. Results: The mutation of p53 gene was present in all three cell lines tested. In PC-9, PC-14 and H69, the altered mobility was detected in exon 7, 7 and 5, respectively. Conclusion: These results suggest that p53 gene mutation plays an important role in certain steps of the carcinogenesis of human non-small cell and small cell lung cancer.

• Korean Journal of Microbiology
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• v.42 no.2
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• pp.89-94
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• 2006

The purpose of this study is to develop the strain-specific PCR primers for the identification of prevotella inter-media ATCC 49046 which is frequently used in the pathogenesis studies of periodontitis. The Hind III-digested genomic DNA of P. intermedia ATCC 49046 were cloned by random cloning method. The specificity of cloned DNA fragments were determined by Southern blot analysis. The nucleotide sequence of cloned DNA probes was determined by chain termination method. The PCR primers were designed based on the nucleotide sequence of cloned DNA fragment. The data showed that Pig6 DNA probe were hybridized with the genomic DNA from P. intermedia strains (ATCC $25611^T$ and 49046) isolated from the Westerns, not the strains isolated from Koreans. The Pig6 DNA probe were consisted of 813 bp. Pig6-F3 and Pig6-R3 primers, designed base on the nucleotide Sequences Of Pig6 DNA Probe, were 3150 specific to the only both P. intermedia ATCC $25611^T$ and P. intermedia ATCC 49046. In the other hand, Pig6-60F and Pig6-770R primers were specific to the only P. intermedia ATCC 49046. The two PCR primer sets could detect as little as 4 pg of chromosomal DNA of P. intermedia. These results indicate that Pig6-60F and Pig6-770R primers have proven useful for the identification of P. intermedia ATCC 49046, especially with regard to the maintenance of the strain.

• Journal of Genetic Medicine
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• v.4 no.1
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• pp.72-79
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• 2007

Purpose : Glycogen storage disease type III (GSD-III) is a rare autosomal recessive disorder of glycogen metabolism. The affected enzyme, amylo-1,6-glucosidase, 4-alpha-glucanotransferase (AGL, glycogen debranching enzyme), is responsible for the debranching of the glycogen molecule during catabolism. The disease shows clinical and biochemical heterogeneity, reflecting genotype-phenotype heterogeneity among different patients. In this study, we aim at analyzing mutations of the AGL gene in three unrelated Korean GSD-III patients, and characterizing their clinical and laboratory findings. Methods : We characterized the clinical features of three unrelated Korean GSD-III patients by biochemical, histological and imaging studies. The 35 exons and part of exon-intron boundaries of AGL were analyzed by direct sequencing using genomic DNA extracted from the peripheral leukocytes of patients. Results : Diverse clinical features were observed in these patients including hepatomegaly (all patients), seizures (patient 2), grow th failure (patients 1 and 2), hyperlipidemia (patients 1 and 3), raised transaminase and creatine kinase concentrations (all patients), and mild cardiomyopathy (patient 2). Liver transplantation w as performed in patient 2 due to progressive hepatic fibrosis. A dministration of uncooked corn starch maintained normoglycemia and improved biochemical and growth profiles. DNA sequence analysis revealed mutations in 5 out of 6 alleles. Patient 1 was a compound heterozygote of c.1282 G>A (p.R428K) and c.1306delA (p.S603PfsX6), patient 2 had c.1510_1511insT (p.Y 504L fsX 10), and patient 3 had c.3416 T >C (p.L 1139P) and c.1735+1 G>T (p.Y 538_R578delfsX 4) mutations. A part from the p.R428K mutation, the 4 other substitutions identified w ere nov el. Conclusion : GSD-III patients display variable phenotypic characteristics resembling those of GSD-Ia. Molecular defects in the AGL gene of Korean GSD-III patients are genetically heterogeneous.

• Horticultural Science & Technology
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• v.28 no.3
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• pp.449-455
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• 2010

'Moulinrouge' was selected as the best regenerating cultivar among 18 different spray-type chrysanthemum cultivars bred in the Gyeongnam Flowers Breeding Research Institute. When the leaf explants from standard- and spray-type chrysanthemum 'Jinba' and 'Moulinrouge' were incubated on MS basal medium supplemented with $0.5mg{\cdot}L^{-1}$ BA and $1.0mg{\cdot}L^{-1}$ NAA, both 'Jinba' and 'Moulinrouge' induced adventitious shoots that can be regenerated into plantlets. Based on these regeneration conditions, we developed an efficient $Agrobacterium$-mediated chrysanthemum 'Moulinrouge' transformation method by using sequential selection of shoots from low ($10mg{\cdot}L^{-1}$) to high ($30mg{\cdot}L^{-1}$) concentrations of kanamycin after co-cultivation of leaf explants with $Agrobacterium$ for 10 days and induction of shoots. All kanamycin resistant plants investigated with genomic PCR analysis carried the report gene, $AtSICKLE$, in their genome. Although expression levels of the report gene in the transgenic plants investigated with RT-PCR were relatively low because of inefficiency of CaMV 35S promoter in chrysanthemum, transgenic lines expressing $AtSICKLE$ efficiently showed leaf epinasty phenotype. We expect that our results will provide a useful method that can perform a high-throughput investigation of genes isolated and studied well in model plants for molecular breeding of chrysanthemum.

• ANNALS OF ANIMAL RESOURCE SCIENCES
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• v.29 no.4
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• pp.150-157
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• 2018

The Zygote arrest 1 (ZAR1) gene is known to affect early embryonic development in various vertebrates. In this study, we performed the association analysis to check whether there is any significant relationship between semen kinematic characteristics and the ZAR1 gene. To determine semen kinematic characteristics, we measured motility (MOT), straight-line velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP), linearity (LIN), straightness (STR), amplitude of lateral head displacement (ALH), and beat cross frequency (BCF) of spermatozoa in boars. In order to detect single nucleotide polymorphisms (SNPs), we extracted genomic DNA from multiple Duroc boars, and then subsequently used them in sequencing reactions. As a result, three SNPs were detected in the intronic region of ZAR1 gene (g.2435T>C in intron 2, g.2605G>A and g.4633A>C in intron 3 ). SNPs g.2435T>C and g.2605G>A were significantly associated with MOT (p<0.01) and VSL (p<0.05), and g.4633A

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.9
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• pp.1204-1209
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• 2004

Random amplification of polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR) analysis was carried out using DNA samples of 30 animals of Rathi cattle and 42 animals of Tharparkar cattle. Genomic DNA was isolated as per standard protocol and evaluated for its quality, purity and concentration. Twenty three random primers were screened out of which 15 primers yielded satisfactory amplifications and were used for further analysis. Average numbers of polymorphic fragments per primer were 7.07${\pm}$0.86 in Rathi and 6.80${\pm}$0.61 in Tharparkar cattle. The percentage of polymorphic bands in these two cattle breeds were 86 and 87%, respectively. Within breed genetic similarities for pooled over primers in the animals of Rathi and Tharparkar breeds were .577${\pm}$0.30 and 0.531${\pm}$0.02, respectively on the basis of band frequency (BF) and 0.645${\pm}$0.04 and 0.534${\pm}$0.04, respectively on the basis of band sharing (BS). Averages of between breed genetic similarities for pooled over primers were 0.97 and 0.92 according to BF and BS, respectively, which reflect higher degree of genetic similarity between Rathi and Tharparkar cattle breeds. Index of genetic distance based on BF and BS for pooled over primers was 0.030${\pm}$0.011 and 0.088${\pm}$0.031, respectively. Percentage of polymorphic bands and within-breed genetic similarities on the basis of band frequency (BF) and band sharing (BS) for pooled over primers revealed higher genetic similarity in Rathi than Tharparkar cattle population. High estimates of between breed genetic similarities for pooled over primers indicated that either Rathi is having decent from Tharparkar or both the cattle breeds are having common descent. Low value of Index of genetic distances between these two cattle breeds may be due to the fact that Rathi and Tharparkar cattle breeds are the native of Thar Desert in Northwest India. The results of between breed genetic distances also confirm the existence of high degree of genetic similarity between these two breeds of cattle.

• Journal of Life Science
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• v.19 no.5
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• pp.566-572
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• 2009

A recombinant plasmid, pDH3, was obtained from the genomic library of Serattia marcescens KCTC 2172, and several recombinant subclones constructed from pDH3. The nucleotide sequence of a 5,137 bp segment, pPH4, was determined and three open reading frames were detected. The three ORFs encoded the phosphate specific transport (pst) operon, which was pstC, pstA, and pstB, with the same direction of transcription. Comparison of the pst operon of S. marcescens with that of other organisms revealed that the genes for pstS and phoU were missing. A potential CRP bonding site and pho box sequence was found in the upstream of the putative promoter at the regulatory region. Analysis of the nucleotide sequence showed that homology in amino acid sequences between the PstC protein and Yersinia sp., Vibrio sp., and Pseudomonas sp. were 49, 37 and 33%, respectively. The PstA protein and Yersinia sp., Vibrio sp., and Pseudomonas sp. showed homologies of 64, 51, and 47%, respectively. PstB protein and Methanocaldococcus sp., E. coli, and Mycoplasma sp. showed homologies of 60, 50, and 48%, respectively. The pst genes could be expressed in vivo and positively regulated by cAMP-CRP. The E. coli strain harboring plasmid pPH7, with pst genes, increased with the transport of phosphate.

• 한국균학회소식:학술대회논문집
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• 2015.05a
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• pp.41-41
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• 2015

BSA-seq technologies, combined Bulked Segregant Analysis (BSA) and Next-Generation Sequencing (NGS), are making it faster and more efficient to establish the association of agronomic traits with molecular markers or candidate genes, which is the requirement for marker-assisted selection in molecular breeding. Clubroot disease, caused by Plasmodiophora brassicae, is a serious threat to Brassica crops. Even we have breed new clubroot resistant varieties of Chinese cabbage (B. rapa ssp. pekinesis), the underlying genetic mechanism is unclear. In this study, an $F_2$ population of 340 plants were inoculated with P. brassicae from Xinye (Pathotype 2 on the differentials of Williams). Resistance phenotype segregation ratio for the populations fit a 3:1 (R:S) segregation model, consistent with a single dominant gene model. Super-BSA, using re-sequencing the parents, extremely R and S DNA pools with each 50 plants, revealed 3 potential candidate regions on the chromosome A03, with the most significant region falling between 24.30 Mb and 24.75 Mb. A linkage map with 31 markers in this region was constructed with several closely linked markers identified. A Major QTL for clubroot resistance, CRq, which was identified with the peak LOD score at 169.3, explaining 89.9% of the phenotypic variation. And we developed a new co-segregated InDel marker BrQ-2. Joint BSA-seq and traditional QTL analysis delimited CRq to an 250 kb genomic region, where four TIR-NBS-LRR genes (Bra019409, Bra019410, Bra019412 and Bra019413) clustered. The CR gene CRq and closely linked markers will be highly useful for breeding new resistant Chinese cabbage cultivars.