• Asian-Australasian Journal of Animal Sciences
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• v.17 no.3
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• pp.309-312
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• 2004

We performed exon trapping in order to locate functional genes on chicken chromosomes (GGA) and to identify functional gene sequences from chicken cosmids. Sequence analysis of 100 clones revealed 17 putative exons, five of which were identified with known sequences in a gene database search: thymopoietin beta (TMPO), U5 snRNP-specific 40 kDa protein (HPRP8BP), dihydropyridine receptor alpha 1 subunit (CACNL1A3), cystein string protein (CPS) and C15orf4. We attempted to map the genes to chicken chromosomes by using FISH and linkage analysis. The chromosomal localizations were GGA1 (TMPO), GGA10 (C15orf4), GGA23 (HPRP8BP) and GGA28 (CPS) by FISH and linkage analysis, while that of CACNL1A3 was predicted to be on a microchromosome by FISH but not by linkage analysis. Comparative mapping analyses between chickens and humans for the genes revealed both known and new synteny. The syntenic conservation between GGA1 and human chromosome (HSA) 12q23 (TMPO) and between GGA10 and HSA15q25 (C15orf4), were consistent with a recent publication, while two new syntenies were observed between GGA28 and HSA20q13.3 in CPS and between GGA23 and HSA1p34-35 in HPRP8BP. The information of presently mapped genes can contribute as anchor markers based on functional genes and the construction of a comparative map.

• Journal of Plant Biology
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• v.37 no.2
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• pp.167-173
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• 1994

Poly(A)+ RNA was selected from Canavalia lineata root nodule RNA through oligo(dT) cellulose column and used for construction of a cDNA library using λgt10-EcoRI arms. The size of the library was 7.2$\times$105 pfu/mL. A full length leghemoglobin (Lb) cDNA clone, pCILb1(687 bp) isolated with soybean Lb probe, contained one open reading frame (ORF) of 447 bp with 54 bp plus 186 bp at 5' and 3' untranslated region, respectively. A consensus sequence of plant translation start region (AAAATGGG) was found at 5' untranslated region, and two polyadenylation-related sequence (AATAAA, AATAAG) and a conserved motif between them (gACTTGTT) were found upstream of poly(A)+ tail consisted of 13 (A)s at 3' untranslated region. The ORF encoded a polypeptide consisted of 149 amino acids with a molecular weight of 16.2 kD. Deduced amino acid sequences showed high degree of homology values with those of other Lbs ranging from 66% (Casuarina glauca) to 85% (Glycine max).

• Journal of Plant Biology
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• v.36 no.4
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• pp.415-423
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• 1993

대두의 uricase II cDNA를 탐침으로 plaque 혼성화 방법에 의해 해녀콩의 뿌리를 cDNA library로부터의 두 개의 phage 클론(λCINUO-01, λCINUO-02)을 선별하였다. 두 phage 클론은 약 1.6 kb와 1.0 kb의 insert를 갖고 있었으며 이들의 염기서열을 결정하기 위하여 pUC19과 pBSKS vector에 subcloing(pcCLNUO-01, pcCLNUO-02)하였다. Sanger법에 의해 염기서열을 결정한 결과, 두 클론은 각각 1,611 bp와 1,024 bp로 이루어져 있었으며 pcCINUO-01은 308개의 아미노산, pcCINUO-02는 301개의 아미노산을 암호화하는 open reading frame(ORF)을 갖고 있었다. 두 클론의 ORF의 염기서열은 대두의 uricase II와 각각 88.9%, 89.3%의 상동성을 보여주었으며, 아미노산 서열은 84.1%, 85.4%의 상동성을 보여주었다. pcCINUO-01의 경우, 종결코돈으로부터 313 NT 하류쪽에 진핵생물의 poly(A) 첨가신호인 AATAAA 서열이 존재하였으며 이로부터 21 NT 하류쪽에 17 잔기의 poly(A)가 존재하였다. 두 클론의 염기서열에서 추정된 아미노산 서열의 카르복시 말단에는 세포질에서 합성된 몇몇 단백질들이 peroxisome으로 수송되는데 필요한 신호서열인 Ser-Lys-Leu-COOH 서열이 존재하고 있었다. 두 클론의 염기서열을 토대로 아미노산 조성을 살펴본 결과, 염기성 아미노산(Arg, His, Lys)과 산성 아미노산(Asp, Glu)이 각각 46 대 35, 47 대 35의 비를 보여주었는데 이는 uricase II 단백질의 염기성 성질을 보여주는 결과로 추정된다. Northern 혼성화 결과 해녀콩에서 uricase II는 뿌리혹에서만 특이적으로 발현됨을 알 수 있었고 게놈 혼성화 반응 결과는 uricase II 유전자가 해녀콩 게놈상에 유전자 가족으로는 존재할 수 있음을 보여주었다.

• Journal of Microbiology and Biotechnology
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• v.33 no.2
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• pp.211-218
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• 2023

A cryptic plasmid (pTH32) was characterized from Tetragenococcus halophilus 32, an isolate from jeotgal, Korean traditional fermented seafood. pTH32 is 3,198 bp in size with G+C content of 35.84%, and contains 4 open reading frames (ORFs). orf1 and orf2 are 456 bp and 273 bp in size, respectively, and their translation products showed 65.16% and 69.35% similarities with RepB family plasmid replication initiators, respectively, suggesting the rolling-circle replication (RCR) mode of pTH32. orf3 and orf4 encodes putative hypothetical protein of 186 and 76 amino acids, respectively. A novel Tetragenococcus-Escherichia coli shuttle vector, pMJ32E (7.3 kb, Em^r), was constructed by ligation of pTH32 with pBluescript II KS(+) and an erythromycin resistance gene (ErmC). pMJ32E successfully replicated in Enterococcus faecalis 29212 and T. halophilus 31 but not in other LAB species. A pepA gene, encoding aminopeptidase A (PepA) from T. halophilus CY54, was successfully expressed in T. halophilus 31 using pMJ32E. The transformant (TF) showed higher PepA activity (49.8 U/mg protein) than T. halophilus 31 cell (control). When T. halophilus 31 TF was subculturd in MRS broth without antibiotic at 48 h intervals, 53.8% of cells retained pMJ32E after 96 h, and only 2.4% of cells retained pMJ32E after 14 days, supporting the RCR mode of pTH32. pMJ32E could be useful for the genetic engineering of Tetragenococcus and Enterococcus species.

• Journal of Veterinary Science
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• v.21 no.6
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• pp.73.1-73.10
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• 2020

Background: Bovine papilloma is a neoplastic disease caused by bovine papillomaviruses (BPVs), which were recently divided into 5 genera and at least 24 genotypes. Objectives: The complete genome sequence of BPV type 15 (BPV Aks-02), a novel putative BPV type from skin samples from infected cows in Southern Xinjiang China, was determined by collecting warty lesions, followed by DNA extraction and amplicon sequencing. Methods: DNA was analyzed initially by polymerase chain reaction (PCR) using the degenerate primers FAP59 and FAP64. The complete genome sequences of the BPV Aks-02 were amplified by PCR using the amplification primers and sequencing primers. Sequence analysis and phylogenetic analysis were performed using bio-informatic software. Results: The nucleotide sequence of the L1 open reading frame (ORF) of BPV Aks-02 was 75% identity to the L1 ORF of BPV-9 reference strain from GenBank. The complete genome consisted of 7,189 base pairs (G + C content of 42.50%) that encoded 5 early (E8, E7, E1, E2, and E4) and 2 late (L1 and L2) genes. The E7 protein contained a consensus CX₂CX₂₉CX₂C zinc-binding domain and a LxCxE motif. Among the different members of this group, the percentages of the complete genome and ORFs (including 5 early and 2 late ORFs) sequence identity of BPV Aks-02 were closer to the genus Xipapillomavirus 1 of the Xipapillomavirus genus. Phylogenetic analysis and sequence similarities based on the L1 ORF of BPV Aks-02 revealed the same cluster. Conclusions: The results suggest that BPV type (BPV Aks-02) clustered with members of the Xipapillomavirus genus as BPV 15 and were closely related to Xipapillomavirus 1.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1995.06b
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• pp.1-19
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• 1995

The plasmid pJEL 101 contains a highly repetitive element from the genome of Xanthomonas oryae pv. oryzae that has properties of an insertional element. The insertional nature of the element, hereto referred to as IS203, was confirmed by molecular analyses of the element and three related elements that were isolated from X. oryzae. The related sequences were isolated on the basis of transposition to the transposon-trapping vector pL3SAC and hybridization with pJEL101. The trapped elements (IS203a, IS203b, and IS203c) were each composed of 1,055 base pairs with 25 base terminal inverted repeats. The elements caused a three base pair target site duplication at the site of insertion in the sacRB gene. The sequence of pJEL 101 has 96% base pair identity with IS203a and 99% identity with IS203a and IS203c but lacks three nucleotides of the consensus left terminal repeat. IS203b has the same DNA sequences as IS203c but is inserted ito the sacRB gene in the opposite orientation. The longest open reading frame of IS203a could code for a protein of 318 amino acids and molecular weight of 37, 151. A search of the Genbank database revealed that IS203 has 51% identity with 909 nucleotides of IS4551 from Escherichia coli. The predicted protein of ORF1 has 40% and 30% amino acid identity to the ORF1 of Tn4551 and the transposase of IS30, respectively.

• Biotechnology and Bioprocess Engineering:BBE
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• v.11 no.4
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• pp.337-343
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• 2006

Trimethylamine dehydrogenase (TMADH, EC 1.5.99.7), an iron-sulfur flavoprotein that catalyzes the oxidative demethylation of trimethylamine to form dimethylamine and formaldehyde, was purified from Methylophaga sp. strain SK1. The active TMADH was purified 12.3-fold through three purification steps. The optimal pH and temperature for enzyme activity was determined to be 8.5 and $55^{\circ}C$, respectively. The $V_{max}\;and\;K_m$ values were 7.9 nmol/min/mg protein and 1.5 mM. A genomic DNA of 2,983 bp from Methylophaga sp. strain SK1 was cloned, and DNA sequencing revealed the open reading frame (ORF) of the gene coding for TMADH. The ORF contained 728 amino acids with extensive identity (82%) to that of Methylophilus methylotrophus $W_3A_1$.

• Korean Journal of Microbiology
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• v.41 no.2
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• pp.99-104
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• 2005

Corynebacterial clones which exert regulatory effects on the expression of the glyoxylate bypass genes were isolated using a reporter plasmid carrying the enteric lacZ fused to the aceB promoter of Corynebacterium glutamicum. Some clones carried common fragments as turned out by DNA mapping technique. Subcloning analysis followed by the measurement of $\beta-galactosidase$ activity in Escherichia coli identified the region responsible for the aceB-repressing activity. Sequence analysis of the DNA fragment identified two independent ORFs of ORF1 and ORF2. Among them, ORF2 was turned out to be responsible for the aceB-repressing activity. ORF1 encoded a 23,216 Da protein composed of 206 amino acids. Sequence similarity search indicated that the ORF may encode a ECF-type $\sigma$ factor and designated sigH. To identify the function of sigH, C. glutamicum sigH mutant was constructed by gene disruption technique and the sigH mutant showed growth retardation as compared to the wild type strain. In addition, the mutant strain showed sensitivity to oxidative-stress generating agent plumbagin. This result imply that sigH is probably involved in the stress response occurring during normal cell growth.

• Microbiology and Biotechnology Letters
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• v.30 no.4
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• pp.312-319
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• 2002

The Sporosarcina psychrophilia pyrB gene, which encodes aspartate transcarbamylase (ATcase), was cloned on Sau3AI restriction endonuclease fragment inserted into pUC19 plasmid vector, S. psychrophilia pyrB gene was expressed in Escherichia coli pyrB mutant for the complementation test. The sequence of 2,606 nucleotides including putative pyrB gene was determined. The region contained one full open reading frame (ORf) and two partial ORFs. The deduced amino acid sequence of the second ORF showed 59% identity with that of Bacillus caldolyticus ATCase. The first and third partial ORFs were closely related to the uracil permease (pyrP) and dihydroorotase (pyrC), respectively. Besides, potential terminator, antiterminator, and anti-antiterminator structures were found in the intergenic region between pyrP and pyrB. These results suggested that S. psychrophilia pyrimidine nucleotide biosynthesis genes are clustered as well as other Bacillus sp. Over-expressed product of pyrB encoding ATCase was purified and analyzed by the SDS-PAGE. The purified PyrB protein turned out to be molecular mass of 27 kDa and showed ATCase activity.

• Journal of Microbiology and Biotechnology
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• v.21 no.3
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• pp.277-283
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• 2011

Glyoxalase I catalyzes the conversion of methylglyoxal to S-D-lactoylglutathione in the presence of glutathione. The structural gene of glyoxalase I (GLO1) was cloned from an osmotolerant yeast, Candida magnoliae, which produces a functional sweetener, erythritol, from sucrose. DNA sequence analysis revealed that the uninterrupted open reading frame (ORF) of C. magnoliae GLO1 (CmGLO1) spans 945 bp, corresponding to 315 amino acid residues, and shares 45.2% amino acid sequence identity to Saccharomyces cerevisiae Glo1. The cloned ORF in a multicopy constitutive expression plasmid complemented the glo1 mutation of S. cerevisiae, confirming that it encodes Glo1 in C. magnoliae. The responses of CmGLO1 to environmental stresses were different from those of S. cerevisiae, which only responds to osmotic stress. An enzyme activity assay and reverse transcription polymerase chain reaction revealed that the expression of CmGLO1 is induced by stress inducers such as methylglyoxal, $H_2O_2$, KCl, and NaCl. The GenBank Accession No. for CmGLO1 is HM000001.