• Title/Summary/Keyword: DNA sequence comparison

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Construction of an Agroinfectious Clone of a Korean Isolate of Sweet Potato Symptomless Virus 1 and Comparison of Its Infectivity According to Agrobacterium tumefaciens Strains in Nicotiana benthamiana

  • Phuong T. Ho;Hee-Seong Byun;Thuy T. B. Vo;Aamir Lal;Sukchan Lee;Eui-Joon Kil
    • The Plant Pathology Journal
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    • v.39 no.3
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    • pp.255-264
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    • 2023
  • Sweet potato symptomless virus 1 (SPSMV-1) is a single-stranded circular DNA virus, belonging to the genus Mastrevirus (family Geminiviridae) that was first identified on sweet potato plants in South Korea in 2012. Although SPSMV-1 does not induce distinct symptoms in sweet potato plants, its co-infection with different sweet potato viruses is highly prevalent, and thus threatens sweet potato production in South Korea. In this study, the complete genome sequence of a Korean isolate of SPSMV-1 was obtained by Sanger sequencing of polymerase chain reaction (PCR) amplicons from sweet potato plants collected in the field (Suwon). An infectious clone of SPSMV-1 (1.1-mer) was constructed, cloned into the plant expression vector pCAMBIA1303, and agro-inoculated into Nicotiana benthamiana using three Agrobacterium tumefaciens strains (GV3101, LBA4404, and EHA105). Although no visual differences were observed between the mock and infected groups, SPSMV-1 accumulation was detected in the roots, stems, and newly produced leaves through PCR. The A. tumefaciens strain LBA4404 was the most effective at transferring the SPSMV-1 genome to N. benthamiana. We confirmed the viral replication in N. benthamiana samples through strand-specific amplification using virion-sense- and complementary-sense-specific primer sets.

Complete Chloroplast Genome assembly and Annotation of Milk Thistle (Silybum marianum) and Phylogenetic Analysis

  • Hwajin Jung;Yedomon Ange Bovys Zoclanclounon;Jeongwoo Lee;Taeho Lee;Jeonggu Kim;Guhwang Park;Keunpyo Lee;Kwanghoon An;Jeehyoung Shim;Joonghyoun Chin;Suyoung Hong
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2022.10a
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    • pp.210-210
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    • 2022
  • Silybum marianum is an annual or biennial plant from the Asteraceae family. It can grow in low-nutrient soil and drought conditions, making it easy to cultivate. From the seed, a specialized plant metabolite called silymarin (flavonolignan complex) is produced and is known to alleviate the liver from hepatitis and toxins damages. To infer the phylogenetic placement of a Korean milk thistle, we conducted a chloroplast assembly and annotation following by a comparison with existing Chinese reference genome (NC_028027). The chloroplast genome structure was highly similar with an assembly size of 152,642 bp, an 153,202 bp for Korean and Chinese milk thistle respectively. Moreover, there were similarities at the gene level, coding sequence (n = 82), transfer RNA (n = 31) and ribosomal RNA (n = 4). From all coding sequences gene set, the phylogenetic tree inference placed the Korean cultivar into the milk thistle clade; corroborating the expected tree. Moreover, an investigation the tree based only on the ycf1 gene confirmed the same tree; suggesting that ycf1 gene is a potential marker for DNA barcoding and population diversity study in milk thistle genus. Overall, the provided data represents a valuable resource for population genomics and species-centered determination since several species have been reported in the Silybum genus.

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Dietary composition of two coexisting bat species, Myotis ikonnikovi and Plecotus ognevi, in the Mt. Jumbong forests, South Korea

  • Sungbae Joo;Injung An;Sun-Sook Kim
    • Journal of Ecology and Environment
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    • v.47 no.4
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    • pp.168-176
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    • 2023
  • Background: Many insectivorous bats have flexible diets, and the difference in prey item consumption among species is one of the key mechanisms that allows for the avoidance of interspecies competition and promotes coexistence within a microhabitat. In Korea, of the 24 bat species that are known to be distributed, eight insectivorous bats use forest areas as both roosting and foraging sites. Here, we aimed to understand the resource partitioning and coexistence strategies between two bat species, Myotis ikonnikovi and Plecotus ognevi, cohabiting the Mt. Jumbong forests, by comparing the differences in dietary consumption based on habitat utilization. Results: Upon examining their dietary composition using the DNA meta-barcoding approach, we identified 403 prey items (amplicon sequence variants). A greater prey diversity including Lepidoptera, Diptera, Coleoptera, and Ephemeroptera, was detected from M. ikonnikovi, whereas most prey items identified from P. ognevi belonged to Lepidoptera. The diversity index of prey items was higher for M. ikonnikovi (H': 5.67, D: 0.995) than that for P. ognevi (H': 4.31, D: 0.985). Pianka's index value was 0.207, indicating little overlap in the dietary composition of these bat species. Our results suggest that M. ikonnikovi has a wider diet composition than P. ognevi. Conclusions: Based on the dietary analysis results, our results suggests the possibility of differences in foraging site preferences or microhabitat utilization between two bat species cohabiting the Mt. Jumbong. In addition, these differences may represent one of the important mechanism in reducing interspecific competition and enabling coexistence between the two bat species. We expected that our results will be valuable for understanding resource partitioning and the coexistence of bats inhabiting the Korean forests.

Comparative Quantitative Study of Surfactant Protein C mRNA by Filter Hybridization and Solution Hybridization in Rats (Filter Hybridization과 Solution Hybridization 방법에 의한 백서 Surfactant Protein C mRNA 정량측정의 비교)

  • Kim, Jin-Ho;Sohn, Jang-Won;Yang, Seok-Chul;Yoon, Ho-Joo;Shin, Dong-Ho;Park, Sung-Soo
    • Tuberculosis and Respiratory Diseases
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    • v.51 no.6
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    • pp.517-529
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    • 2001
  • Background : Surfactant protein C(SP-C) is a hydrophobic 5,000 dalton molecule. SP-C has the primary roles in accelerating surface spreading of a surfactant phospholipid. The filter hybridization and solution hybridization assays are both rapid and sensitive and can be used to measure the RNAs complementary to any cloned DNA sequence. Methods : The authors measured the SP-C mRNA levels quantitatively using solution hybridization and filter hybridization assays to obtain a standard curve equation to quantify the mRNA of unknown samples comparatively. Results : 1. The minimum level of the specimens by solution hybridization was 3 pg for SP-C mRNA. 2. The standard curve equation of the solution hybridization assay between the counts per minute(Y) and the SP-C mRNA transcript input(X) was Y=6.46 X+244. The correlation coefficient was 0.99. 3. The minimum detection level of specimens by filter hybridization was 0.1 ng for SP-C mRNA. 4. The standard curve equation of the filter hybridization assay between the counts per minute(Y) and SP-C mRNA transcript input(X) is Y=2541.6 X+252.7. The correlation coefficient was 0.99. Conclusions : A comparison of CPM/filter in the linear range allowed an accurate and reproducible estimation of the SP-C mRNA copy number. Filter hybridization and solution hybridization assays are both rapid and sensitive and can be used to measure the RNAs complementary to any cloned DNA sequence. It is ideally suited to situations where accurate quantitation of multiple samples is required.

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Cloning and Expression of FSHb Gene and the Effect of $FSH{\beta}$ on the mRNA Levels of FSHR in the Local Chicken

  • Zhao, L.H.;Chen, J.L.;Xu, H.;Liu, J.W.;Xu, Ri Fu
    • Asian-Australasian Journal of Animal Sciences
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    • v.23 no.3
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    • pp.292-301
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    • 2010
  • Follicle-stimulating hormone (FSH) is a pituitary glycoprotein hormone that is encoded by separate alpha- and betasubunit genes. It plays a key role in stimulating and regulating ovarian follicular development and egg production in chicken. FSH signal transduction is mediated by the FSH receptor (FSHR) that exclusively interacts with the beta-subunit of FSH, but characterization of prokaryotic expression of the FSHb gene and its effect on the expression of the FSHR gene in local chickens have received very little attention. In the current study, the cDNA fragment of the FSHb gene from Dagu chicken was amplified using reverse transcription polymerase chain reaction (RT-PCR), and inserted into the pET-28a (+) vector to construct the pET-28a-FSHb plasmid. After expression of the plasmid in E. coli BL21 (DE3) under inducing conditions, the recombination protein, $FSH{\beta}$ subunit, was purified and injected into the experimental hens and the effect on the mRNA expression levels of the FSHR gene was investigated. Sequence comparison showed that the coding region of the FSHb gene in the local chicken shared 99%-100% homology to published nucleotides in chickens; only one synonymous nucleotide substitution was detected in the region. The encoded amino acids were completely identical with the reported sequence, which confirmed that the sequences of the chicken FSHb gene and the peptides of the $FSH{\beta}$ subunit are highly conserved. This may be due to the critical role of the normal function of the FSHb gene in hormonal specificity and regulation of reproduction. The results of gene expression revealed that a recombinant protein with a molecular weight of about 19 kDa was efficiently expressed and it was identified by Western blotting analysis. After administration of the purified $FSH{\beta}$ protein, significantly higher expression levels were demonstrated in uterus, ovary and oviduct samples (p<0.05). These observations suggested that the expressed $FSH{\beta}$ protein possesses biological activity, and has a potential role in regulation of reproductive physiology in chickens.

Isolation and Characterization of a Novel Agar-Degrading Marine Bacterium, Gayadomonas joobiniege gen, nov, sp. nov., from the Southern Sea, Korea

  • Chi, Won-Jae;Park, Jae-Seon;Kwak, Min-Jung;Kim, Jihyun F.;Chang, Yong-Keun;Hong, Soon-Kwang
    • Journal of Microbiology and Biotechnology
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    • v.23 no.11
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    • pp.1509-1518
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    • 2013
  • An agar-degrading bacterium, designated as strain $G7^T$, was isolated from a coastal seawater sample from Gaya Island (Gayado in Korean), Republic of Korea. The isolated strain $G7^T$ is gram-negative, rod shaped, aerobic, non-motile, and non-pigmented. A similarity search based on its 16S rRNA gene sequence revealed that it shares 95.5%, 90.6%, and 90.0% similarity with the 16S rRNA gene sequences of Catenovulum agarivorans $YM01^T$, Algicola sagamiensis, and Bowmanella pacifica W3-$3A^T$, respectively. Phylogenetic analyses demonstrated that strain $G7^T$ formed a distinct monophyletic clade closely related to species of the family Alteromonadaceae in the Alteromonas-like Gammaproteobacteria. The G+C content of strain $G7^T$ was 41.12 mol%. The DNA-DNA hybridization value between strain $G7^T$ and the phylogenetically closest strain $YM01^T$ was 19.63%. The genomes of $G7^T$ and $YM01^T$ had an average ANIb value of 70.00%. The predominant isoprenoid quinone of this particular strain was ubiquinone-8, whereas that of C. agarivorans $YM01^T$ was menaquinone-7. The major fatty acids of strain $G7^T$ were Iso-$C_{15:0}$ (41.47%), Anteiso-$C_{15:0}$ (22.99%), and $C_{16:1}{\omega}7c/iso-C_{15:0}2-OH$ (8.85%), which were quite different from those of $YM01^T$. Comparison of the phenotypic characteristics related to carbon utilization, enzyme production, and susceptibility to antibiotics also demonstrated that strain $G7^T$ is distinct from C. agarivorans $YM01^T$. Based on its phenotypic, chemotaxonomic, and phylogenetic distinctiveness, strain $G7^T$ was considered a novel genus and species in the Gammaproteobacteria, for which the name Gayadomonas joobiniege gen. nov. sp. nov. (ATCC BAA-2321 = $DSM25250^T=KCTC23721^T$) is proposed.

Characterization and Expression of Penaeidin 3-2 from Fleshy Prawn Fenneropenaeus chinensis (대하 Penaeidin 3-2 유전자의 동정 및 발현)

  • Park, Eun-Mi;Cho, Hyun Kook;Hong, Gyeong-Eun;Nam, Bo-Hye;Kim, Young-Ok;Kim, Woo-Jin;Lee, Sang-Jun;Han, Hyon Sob;Lee, Jae Yong;Kim, Jong-Sheek;Jang, In-Kwon;Cheong, JaeHun;Choi, Tae-Jin;Kong, Hee Jeong
    • Journal of Marine Bioscience and Biotechnology
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    • v.2 no.1
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    • pp.34-39
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    • 2007
  • Penaeidins are members of a special family of antimicrobial peptides existing in several species of shrimp and play a crucial role in the immunological defense of shrimp. In this study, we isolated and characterized one EST clone (penaeidin) from cDNA library of fleshy prawn Fenneropenaeus chinensis hemocytes. Amino acids sequence comparison and phylogenetic analysis with other known penaeidins revealed that our clone was completely identical to F. chinensis Penaeidin 3-2 (Accession no. ABC33920), which composed of 71 amino acids with a putative signal peptide (1-19) and a cysteine-rich domain (C-terminal part). The expression and distribution of Penaeidin 3-2 transcripts in shrimp were detected in hemocytes, hepatopancreas, and muscles, and that Penaeidin 3-2 was constitutively expressed mainly in hemocytes. The artificial infection of white spot syndrome virus to F. chinensis resulted in Penaeidin 3-2 mRNA up-regulation in hemocytes, suggesting that the possible role of Penaeidin 3-2 in host defense system of F. chinensis.

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EXPRESSION OF DOMINANT NEGATIVE p63 ISOFORM IN WELL-DIFFERENCIATED ORAL SQUAMOUS CELL CARCINOMA (분화도 좋은 구강 편평상피세포암종에서 Dominant Negative p63 isoform의 발현)

  • Kim, In-Su;Kim, Chul-Hwan;Kim, Kyung-Wook
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.33 no.3
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    • pp.191-198
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    • 2007
  • The p53 which is well known as tumor suppressor gene is located at 17p13. p53 is a sequence-specific DNA binding transcription factor that responds to certain cytotoxic stresses, such as DNA damage, by enhancing the transcription of genes that regulate cell-cycle progression as well as programmed cell death. The p63 gene that is located at 3q27-29, is recognized members of the p53 family, and responsible for the transcription of 6 isoforms. Three isoforms ($TAp63{\alpha}$, $TAp63{\beta}$, $TAp63{\gamma}$) contain an N-terminal transactivation (TA) domain and can induce apoptosis. The other 3 isoforms (${\Delta}Np63{\alpha}$, ${\Delta}Np63{\beta}$, ${\Delta}Np63{\gamma}$) lack the TA domain and may function in a dominant-negative fashion by inhibiting the transactivation functions of p53 and TAp63 proteins, and thus act as oncoproteins. A number of studies have investigated the role of p63 in human squamous cell carcinomas from different organs. Only a few studies have examined ${\Delta}Np63$ isoform in oral squamous cell carcinoma including normal epithelium. This study aimed to evaluate expression of ${\Delta}Np63$ isoform in human oral squamous cell carcinoma tissue and normal mucosa. The 3 cases of well differenciated oral squamous cell carcinoma specimen including adjacent normal mucosa were examined, and immunohistochemical study with monoclonal antibody(4A4) and tumor cell apoptosis analysis with Transmission Electon Microscopy were studied. And, RT-PCR analysis was done for expression of ${\Delta}Np63$ isoform. The results were as followed. 1. Normal gingiva showed the restricted p63 expression in basal cell layer. 2. Well differentiated squamous cell carcinoma showed mainly p63 expression in overall area of malignancy, especially in basal cell layer to adjacent stromal tissue. 3. Tumor cells around keratinized area with no p63 expression disclosed less micro-organelle in decreased size cytoplasm and severe chromatin margination with nuclear destruction that means apoptosis. 4. Comparison of mRNA expression of ${\Delta}Np63$ isoform by RT-PCR showed variable expression of ${\Delta}Np63$ isoform, but ${\Delta}Np63{\alpha}$ was most highly expressed in all 3 tumor specimen. From theses results, it should be suggested that ${\Delta}Np63$ isoform expression in well differentiated squamous cell carcinoma was closely related to tumor oncogenesis, expecially overexpression of ${\Delta}Np63{\alpha}$ is a most important factor in tumor genesis of oral squamous cell carcinoma.

Comparison of Immunohistochemical Expression of CBP(cAMP-responsive Element Binding Protein) Transcriptional Co-activator between Premalignant Lesions and Squamous Cell Carcinomas in the Lungs (전암성 폐병변 및 편평상피세포폐암 조직에서 CBP(cAMP-responsive Ele-ment Binding Protein) 전사 공동 활성인자의 면역조직화학적 발현양상의 비교)

  • Shin, Jong Wook;Kim, Jin Soo;Kim, Mi Kyung
    • Tuberculosis and Respiratory Diseases
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    • v.63 no.2
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    • pp.165-172
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    • 2007
  • Background: The pathogenesis of lung cancer includes the accumulation of multiple genetic abnormalities. The CREB-binding protein(CBP) is one of several transcriptional co-activators among various sequence-specific DNA-binding transcription factors. CBP is involved in a wide range of cellular activities, such as DNA repair, cell growth, differentiation, and apoptosis that are suspected of contributing to tumorigenesis. The goal of this study was to evaluate CBP expression in a series of human lung tissues containing normal epithelium, premalignant lesions(hyperplasia and dysplasia) and squamous cell carcinomas. Materials and Methods: Immunohistochemical staining was performed on formalin-fixed paraffin-embedded sections by use of a monoclonal anti-CBP antibody. CBP expression was compared in samples from 120 patients with premalignant and malignant histological types including 20 metaplastic specimens, 40 dysplastic specimens, and 60 squamous cell carcinomas in the lung. Results: CBP expression was seen in 35% (7/20) of the metaplastic specimens. 65% (26/40) of the dysplastic specimens, and 70% (42/60) of the squamous cell carcinomas (p<0.05). According to celluar atypism, CBP expression was 50% (10/20) of the low-grade dysplastic specimens and 80% (16/20) of the high-grade dysplastic specimens(p <0.01). By cellular differentiation, CBP expression was seen in 95% (19/20) of the well differentiated squamous cell carcinomas, 85% (17/20) of the moderately differentiated carcinomas and 30% (6/20) of the poorly differentiated lesions (p <0.05). Conclusion: These results suggest that CBP may have an important role in malignant transformation of precancerous lung lesions and may be a marker for malignancy.

Phylogenetic Analysis of Carassius auratus and C. cuvieri in Lake Yedang Based on Variations of Mitochondrial CYTB Gene Sequences (예당호 붕어와 떡붕어의 CYTB 유전자를 이용한 유연관계 분석)

  • Kim, Gye-Woong;Joe, Sung-Duck;Kim, Hack-Youn;Park, Hee-Bok
    • Journal of Life Science
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    • v.30 no.12
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    • pp.1063-1069
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    • 2020
  • Two crucian carp species (Carassius auratus and C. cuvieri) inhabit Lake Yedang in South Korea, and C. auratus is known to be native to Korea. Classification of these two freshwater fish species is often confused because of their morphological similarity. To distinguish the two species, we conducted phylogenetic and population genetic analyses of C. auratus and C. cuvieri based on their mitochondrial DNA sequences of the cytochrome b gene (CYTB). We also compared our partial CYTB sequence (<1,056 bp) with 10 Chinese, nine Japanese, and two Russian crucian carp fishes. The results of our phylogenetic analysis showed that C. auratus and C. cuvieri were clearly divided into two phylogroups. The nucleotide diversity (π) of C. auratus from Korea, China, and Japan showed a range of 0.146%~0.421%, while the range of π of C. cuvieri from Korea and Japan was lower than those of C. auratus (0.0%~0.054%). Moreover, the comparison of CYTB divergence among crucian carp fishes in China, Japan, and Korea indicated that Korean Carassius fishes were distantly related to those from China and Japan, with two exceptions: the pairwise Fst value between Korean C. auratus and northern Chinese C. auratus was not significantly different. In addition, no significant genetic divergence between Korean and Japanese C. cuvieri was detected. We conclude that, despite the morphological similarities, C. auratus and C. cuvieri should be considered as separate freshwater fish resources in conservation efforts for genetic diversity.