• Journal of Veterinary Science
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• 제22권4호
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• pp.43.1-43.10
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• 2021

Background: The H5 avian influenza viruses (AIVs) of clade 2.3.4.4 circulate in wild and domestic birds worldwide. In 2017, nine strains of H5N6 AIVs were isolated from aquatic poultry in Xinjiang, Northwest China. Objectives: This study aimed to analyze the origin, reassortment, and mutations of the AIV isolates. Methods: AIVs were isolated from oropharyngeal and cloacal swabs of poultry. Identification was accomplished by inoculating isolates into embryonated chicken eggs and performing hemagglutination tests and reverse transcription polymerase chain reaction (RT-PCR). The viral genomes were amplified with RT-PCR and then sequenced. The sequence alignment, phylogenetic, and molecular characteristic analyses were performed by using bioinformatic software. Results: Nine isolates originated from the same ancestor. The viral HA gene belonged to clade 2.3.4.4B, while the NA gene had a close phylogenetic relationship with the 2.3.4.4C H5N6 highly pathogenic avian influenza viruses (HPAIVs) isolated from shoveler ducks in Ningxia in 2015. The NP gene was grouped into an independent subcluster within the 2.3.4.4B H5N8 AIVs, and the remaining six genes all had close phylogenetic relationships with the 2.3.4.4B H5N8 HPAIVs isolated from the wild birds in China, Egypt, Uganda, Cameroon, and India in 2016-2017, Multiple basic amino acid residues associated with HPAIVs were located adjacent to the cleavage site of the HA protein. The nine isolates comprised reassortant 2.3.4.4B HPAIVs originating from 2.3.4.4B H5N8 and 2.3.4.4C H5N6 viruses in wild birds. Conclusions: These results suggest that the Northern Tianshan Mountain wetlands in Xinjiang may have a key role in AIVs disseminating from Central China to the Eurasian continent and East African.

• The Plant Pathology Journal
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• 제35권4호
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• pp.381-388
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• 2019

For several years, temperatures in the Korean peninsula have gradually increased due to climate change, resulting in a changing environment for growth of crops and vegetables. An associated consequence is that emerging species of insect vector have caused increased viral transmission. In Jeju Island, Korea, occurrences of viral disease have increased. Here, we report characterization of five newly collected turnip mosaic virus (TuMV) isolates named KBJ1, KBJ2, KBJ3, KBJ4 and KBJ5 from a survey on Jeju Island in 2017. Full-length cDNAs of each isolate were cloned into the pJY vector downstream of cauliflower mosaic virus 35S and bacteriophage T7 RNA polymerase promoters. Their fulllength sequences share 98.9-99.9% nucleotide sequence identity and were most closely related to previously reported Korean TuMV isolates. All isolates belonged to the BR group and infected both Chinese cabbage and radish. Four isolates induced very mild symptoms in Nicotiana benthamiana but KBJ5 induced a hypersensitive response. Symptom differences may result from three amino acid differences uniquely present in KBJ5; Gly(382)Asp, Ile(891)Val, and Lys(2522)Glu in P1, P3, and NIb, respectively.

• Asian-Australasian Journal of Animal Sciences
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• 제32권5호
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• pp.614-628
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• 2019

Objective: The inhibitory leukocyte immunoglobulin-like receptors (LILRBs) play an important role in innate immunity. The present study represents the first description of the cloning and structural and functional analysis of LILRB1 and LILRB3 isolated from two genetically disparate chicken lines. Methods: Chicken LILRB1-3 genes were identified by bioinformatics approach. Expression studies were performed by transfection, quantitative polymerase chain reaction. Signal transduction was analyzed by western blots, immunoprecipitation and flow cytometric. Cytokine levels were determined by enzyme-linked immunosorbent assay. Results: Amino acid homology and phylogenetic analyses showed that the homologies of LILRB1 and LILRB3 in the chicken line 6.3 to those proteins in the chicken line 7.2 ranged between 97%-99%, while homologies between chicken and mammal proteins ranged between 13%-19%, and 13%-69%, respectively. Our findings indicate that LILRB1 and LILRB3 subdivided into two groups based on the immunoreceptor tyrosine-based inhibitory motifs (ITIM) present in the transmembrane domain. Chicken line 6.3 has two ITIM motifs of the sequence LxYxxL and SxYxxV while line 7.2 has two ITIM motifs of the sequences LxYxxL and LxYxxV. These motifs bind to SHP-2 (protein tyrosine phosphatase, non-receptor type 11) that plays a regulatory role in immune functions. Moreover, our data indicate that LILRB1 and LILRB3 associated with and activated major histocompatibility complex (MHC) class I and ${\beta}2-microglobulin$ and induced the expression of transporters associated with antigen processing, which are essential for MHC class I antigen presentation. This suggests that LILRB1 and LILRB3 are transcriptional regulators, modulating the expression of components in the MHC class I pathway and thereby regulating immune responses. Furthermore, LILRB1 and LILRB3 activated Janus kinase2/tyrosine kinase 2 (JAK2/TYK2); signal transducer and activator of transcription1/3 (STAT1/3), and suppressor of cytokine signaling 1 genes expressed in Macrophage (HD11) cells, which induced Th1, Th2, and Th17 cytokines. Conclusion: These data indicate that LILRB1 and LILRB3 are innate immune receptors associated with SHP-2, MHC class I, ${\beta}2-microglobulin$, and they activate the Janus kinase/signal transducer and activator of transcription signaling pathway. Thus, our study provides novel insights into the regulation of immunity and immunopathology.

• Parasites, Hosts and Diseases
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• 제58권6호
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• pp.709-714
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• 2020

Knockdown resistance (kdr) mutations in the voltage-gated sodium channel (VGSC) of mosquitoes confer resistance to insecticides. Although insecticide resistance has been suspected to be widespread in the natural population of Aedes aegypti in Myanmar, only limited information is currently available. The overall prevalence and distribution of kdr mutations was analyzed in Ae. aegypti from Mandalay areas, Myanmar. Sequence analysis of the VGSC in Ae. aegypti from Myanmar revealed amino acid mutations at 13 and 11 positions in domains II and III of VGSC, respectively. High frequencies of S989P (68.6%), V1016G (73.5%), and F1534C (40.1%) were found in domains II and III. T1520I was also found, but the frequency was low (8.1%). The frequency of S989P/V1016G was high (55.0%), and the frequencies of V1016G/F1534C and S989P/V1016G/F1534C were also high at 30.1% and 23.5%, respectively. Novel mutations in domain II (L963Q, M976I, V977A, M994T, L995F, V996M/A, D998N, V999A, N1013D, and F1020S) and domain III (K1514R, Y1523H, V1529A, F1534L, F1537S, V1546A, F1551S, G1581D, and K1584R) were also identified. These results collectively suggest that high frequencies of kdr mutations were identified in Myanmar Ae. aegypti, indicating a high level of insecticide resistance.

• 한국수산과학회지
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• 제55권5호
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• pp.612-624
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• 2022

A novel Gram-positive, motile, non-spore forming aerobic marine bacterium, designated 107-1^T was isolated from tidal mud collected in Gyehwa-do, South Korea. Cells of strain 107-1^T were short rod or coccoid, oxidase negative, catalase positive and grew at 10-40℃ (with optimum growth at 25-30℃). It utilized menaquinones MK-7 and 8 as its respiratory quinones and its major fatty acids were anteiso-C_15:0 (37.9%), iso-C_16:0 (14.9%), and iso-C_14:0 (10.8%). Phylogenetic analysis based on 16S rRNA gene sequences revealed a distinct clade containing strain 107-1^T and close species Planococcus ruber CW1^T(98.52% sequence similarity), P. faecalis KCTC 33580^T(97.67%), P. kocurii ATCC 43650T(97.48%), P. donghaensis DSM 22276T(97.47%), and P. halocryophilus DSM 24743T(97.37%). Strain 107-1^T contains one circular chromosome (3,513,248bp in length) with G+C content of 44.6 mol%. Estimated ranges for genome to genome distance, average nucleotide identity, and average amino acid identity comparing strain 107-1^T with close taxa were 20.3-34.8%, 77.9-86.9%, and 73.6-92.8%, respectively. Based on polyphasic analysis, strain 107-1^T represents a novel species belonging to the genus Planococcus.

• 한국버섯학회지
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• 제20권3호
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• pp.178-182
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• 2022

버섯의 오랜 역사에도 불구하고 버섯의 유전적 기능과 분자유전학을 응용한 신품종 개발에 대한 연구는 크게 부족한 상황이다. 그러나 최근 유전자 가위인 CRISPR/Cas를 이용한 새로운 유전자 교정 기술이 개발됨에 따라 버섯 연구에서 이 기술을 이용한 다양한 시도가 이루어지고 있다. 특히 선택의 용이성을 위해 외래 유전자 삽입 없이도 고효율로 유전자 편집이 가능한 RNPs를 활용한 연구가 활발히 진행되고 있다. 그러나 RNPs는 원형질체의 세포막을 통과하기에 Cas9이 너무 거대하고 guide RNA가 쉽게 파괴된다는 단점을 가지고 있다. 이러한 단점을 극복하기 위하여 세포막 통과에 용이한 미네랄 성분인 CaP와 PAA를 조합하여 Nanoparticle을 형성함으로써 극복하고자 했다. 표고버섯 단핵 균주인 산조705-13을 이용하여 원형질체 분리에 적합한 Osmotic buffer를 찾기 위하여 0.6M과 1.2M의 Sucrose, Sorbitol, Mannitol, KCl을 처리하였고 그 결과 0.6M Sucrose가 가장 적합한 osmotic buffer임을 확인하였다. 또한 CaP으로 RNPs와 Nanoparticle 복합체를 형성하고 이 복합체가 RNase A로부터 RNPs의 기능을 온전히 보호하는 것을 확인할 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제33권10호
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• pp.1292-1298
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• 2023

PAMB 00755^T, a bacterial strain, was isolated from Korean fir leaves. The strain exhibits yellow colonies and consists of Gram-negative, non-motile, short rods or ovoid-shaped cells. It displays optimal growth conditions at 20℃, 0% NaCl, and pH 6.0. Results of 16S rRNA gene-based phylogenetic analyses showed that strain PAMB 00755^T was most closely related to Sphingomonas chungangi MAH-6^T (97.7%) and Sphingomonas polyaromaticivorans B2-7^T (97.4%), and ≤96.5% sequence similarity to other members of the genus Sphingomonas. The values of average nucleotide identity (79.9-81.3%), average amino acid identity (73.3-75.9%), and digital DNA-DNA hybridization (73.3-75.9%) were significantly lower than the threshold values for species boundaries; these overall genome-related indexes (OGRI) analyses indicated that the strain represents a novel species. Genomic analysis revealed that the strain has a 4.4-Mbp genome encoding 4,083 functional genes, while the DNA G+C content of the whole genome is 66.1%. The genome of strain PAMB 00755^T showed a putative carotenoid biosynthetic cluster responsible for its antioxidant activity. The respiratory quinone was identified as ubiquinone 10 (Q-10), while the major fatty acids in the profile were identified as C_18:1ω7c and/or C_18:1ω6c (summed feature 8). The major polar lipids of strain PAMB 00755^T were diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, and phosphatidylcholine. Based on a comprehensive analysis of genomic, phenotypic, and chemotaxonomic characteristics, we proposed the name Sphingomonas abietis sp. nov. for this novel species, with PAMB 00755^T as the type strain (= KCTC 92781^T = GDMCC 1.3779^T).

• 한국미생물·생명공학회지
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• 제44권3호
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• pp.355-362
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• 2016

수해양성 병원성 미생물로 알려진 Vibrio anguillarum으로부터 mannose-1-phosphate를 mannose-6-phosphate, glucose-1-phosphate를 glucose-6-phosphate로 가역적으로 변환시키는 phosphomannomutase/phosphoglucomutase (pmm/pgm)의 유전자를 sequencing하여 1338 bp의 open reading frame (ORF)을 밝혔다. 이는 446개의 아미노산을 포함하며 47,625 Da을 가지고 있다. 보고된 다른 Vibrio sp.의 pmm/pgm 유전자와 상동성을 비교하였을 때 V. mimicus V. vulnificus, V. splendidus, V. harveyi와 92.3%, 91.4%, 89.9%, 89.9%에 해당하는 상동성을 지니고 있었다. 증폭된 목적 유전자를 pET-28a(+) vector에 연결하여 대장균에서 단백질의 대량발현을 유도하였으며 이는 주로 soluble한 상태로 나왔다. Soluble fraction을 Ni-NTA column chromatography로 정제하여 약 50 kDa의 단백질을 얻었고 이는 주로 mannose-1-phosphate를 이용하는 효소로 확인되었으며 Mg²⁺ 이온이 존재할 때 효소의 활성이 나타나는 것을 확인할 수 있었다. 본 연구의 유전자는 낮은 온도의 stress하에서 발현이 증가됨을 Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)을 통해 확인하였고, 상동성 재조합 (homologous recombination)에 의한 돌연변이 균주 제작을 통해 PMM/PGM protein과 lipopolysaccharide (LPS)의 생합성과의 관계를 규명하였다. V. anguillarum wild type과 mutant로부터 LPS를 분리하였고 sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE)후 silver staining을 통해 LPS의 high molecular weight (HMW) 부분인 O-antigen에서의 변화를 확인하였다. 또한 V. anguillarum wild type과 mutant의 growth와 viability를 확인한 결과 mutant가 wild type보다 정지기까지 더 낮은 생육을 보였으며 viability가 감소함을 확인하였다. 본 연구를 통하여 V. anguillarum의 pmm/pgm 유전자가 미생물의 생육과 LPS 생합성에 관여하고 있음을 알 수 있었다.

• Journal of Plant Biotechnology
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• 제32권3호
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• pp.167-173
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• 2005

본 연구는 고추열매의 capsaicinoid 생합성 조절 기작을 연구하고자 general phenylpropanoid pathway의 2번째 단계에 작용하는 것으로 알려진 3종류의 c4h cDNA clone을 확보하여 염기서열을 분석한 결과, pc4h1와 pc4h2의 크기는 각각 1,775 bp와 1,655 bp으로 505개의 아미노산으로 구성된 펩티드를 암호하는 full length의 ORF를 갖추고 있었으나 pc4h3는 5'-말단의 coding 영역 일부가 소실 되었다. 이들은 공히 모든 cytochrome P450 효소에서 진화학적으로 보존되어 있는 3종류의 conserved region 즉 domain 1(proline-rich region), domain 2 (threonine-containing binding pocket for the oxygen molecule), 그리고domain 3(heme binding region)을 포함하고 있었으며 evolutionary tree분석을 통하여 pc4h1와 pc4h2는 모두 Class I에 속하는 것으로 서로 간에는 95.8%의 유사성을 나타내어 거의 동일한 유전자인 것으로 조사되었다. 특히 Class II로 분류된 Citrus sinensis C4H1과 Phaseolus vulgaris C4H와는 단지 64.9에서 64.5%의 homology를 나타내었다. 또한 pc4h2는 상처를 주지 않은 조직에서는 비교적 적은 양의 mRNA 발현수준을 보였으나 상처를 가한 후 6시간의 열매에서는 400%, 뿌리에서는 200%까지 그 발현 양이 증가하였다. 이와 유사하게 pc4h1의 전사량도 상처에 의하여 유도는 되나 basal level이 pc4h2보다 높아서 발현증가 비율은 그다지 높지 않았다. 반면에 pc4h3 유전자는 상처를 주지 않은 모든 조직에서 거의 발현되지 않았으며 상처에 의하여서도 전혀 반응을 하지 않았다.

• Journal of Plant Biotechnology
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• 제35권3호
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• pp.177-183
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• 2008

본 연구는 동진벼 유래의 Ac/Ds 삽입변이집단의 GUS 분석을 통하여 뿌리 및 미숙종자에서 강하게 GUS가 발현한 개체를 선발하여 FST(flanking sequence tag) 분석 한 결과 Ds 전이 인자가 3번 염색체 zinc finger RING-H2 관련 Oszinc626 유전자의 첫 번째 exon 부위에 single copy로 삽입되어 있었으며, 선발변이체는 뿌리 및 종자 발달이 정상인 동진벼에 비해 매우 낮은 것으로 나타났다. Oszinc626 유전자는 RING-H2 type(C3-H2-C3)으로 $Cys-X_2-Cys-X_{28}-Cys-X-His-X_2-His-X_2-Cys-X_{14}-Cys-X_2-Cys$ 배열이 C-terminus 가장 말단에 위치하며, 49 kDa의 분자량을 가지고 있다. 또한 Southern blot 분석에서 Oszinc626 유전자는 벼 게놈상에 single copy로 존재하였다. RT-PCR을 통한 돌연변이 유전자의 발현분석 결과 250 mM의 염과, $4^{\circ}C$ 저온등과 같은abiotic stress에 의해 발현이 증가함을 보였고, 호르몬처리에 있어서 ABA와 IAA의 식물호르몬을 처리했을 경우 24시간까지 계속해서 발현양이 증가하는 것을 보이는 반면, 2,4-D 처리의 경우 30분 후에 발현이 일시적으로 증가되었으나 이후 발현이 급속히 감소한 것을 보였다. 벼의 조직 별 발현 검정에서 미성숙한 종자, 뿌리 분열조직 및 신초 등 주로 생장점 부위에서 강하게 발현되는 것을 보임에 따라 Oszinc626 유전자의 경우 식물의 생장에 관여하는 주동 유전자의 하나로 판단된다.