• Journal of the Korean Society of Tobacco Science
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• v.21 no.1
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• pp.57-63
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• 1999

Pokeweed antiviral protein II (PAP-II) encoding cDNA was synthesized by reverse-transcriptase polymerase chain reaction (RT-PCR) from Phytolacca american a leaf. The PAP-II cDNA fragment of 974bp was subcloned to pBluescript II SK- SmaI site and the inserted PAP-II cDNA fragment was sequenced by dideoxy sequencing method. The number of nucleotides of PAP-II cDNA coding region containing start and stop codon was 933bp. To develop a virus-resistant tobacco plant, PAP-II cDNA fragment was inserted to pKGT101B and the insertion of PAP-II cDNA fragment was confirmed by restriction enzyme analysis and colony PCR.

• Korean Journal of Pharmacognosy
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• v.10 no.2
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• pp.55-59
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• 1979

Mass spectra of the dammarane triterpenes having open side chain and $C_{20}-C_{25}-epoxy$ side chain were measured. Principal fragment ions were assigned and plausible mechanisms for the formation of the fragment ions were proposed. In general, the triter-penoids of $C_{20}-C_{25}-epoxy$ side chain. produce $h_{1}-species$ fragment ions by the deletion of side chain at $C_{20}-C_{22}$ bond and open side chain triterpenoids produce $h_{2}$ species fragmentions whose mass numbers are higher by two mass unit than those of $h_{1}$ species. The mass number of h species fragment ions will serve as the diagnostic tool for the elucidation of side chain structure of tetracyclic triterpenoids.

• Proceedings of the Korean Society of Food Hygiene and Safety Conference
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• 2002.05a
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• pp.215-215
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• 2002

Catalytic subunit of pyruvate dehydrogenase phosphatase (PDPc) has been suggested to have three major funational domains such as dihydrplipoamide adetyltransferase(E2)-binding domain, regulatory subunit of PDP(PDP)r-binding domain, and calcium-binding domain. In order to identify functional domains, recombinant catalytic subunit of pyruvate dehydrogenase phosphatase(rPDPc) was expressed in E. coli JM101 and purified to near homogeneity using the unique property of PDPc: PDPc binds to the inner lipoyl domain (L2) of E2 of ppyruvate dehydrogenase complex (PDC) in the presence of Ca+2, not under EGTA. PDPc was limited-proteolysed by typsin, chymotypsin, Arg-C, and elastase at pH 7.0 and 30C and N-terminal analysis of the fragments was done. Chymotrypsin, trypsin, and elastase made two major fragments: N-terminal large fragment, approx. 50kD and C-terminal small fragment, approx.10 kDa. Arg-C made three major fragments: N-terminal fragment, approx. 35kD, and central fragment, approx. 15 kD, and C-terminal fragment, approx. 10 kD. This study strongly suggest that PDPc consists of three major functional domains. However, further study should be necessary to identify the functional role.

• Proceedings of the Korean Society of Applied Pharmacology
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• 2002.07a
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• pp.236-236
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• 2002

Pyruvate dehydrogenase phosphatase (PDP) is a mitochondrial protein serine/threonine phosphatase that catalyzes the dephosphorylation and concomitant reactivation of the pyruvate dehydrogenase componant of the pyruvate dehydrogenase complex (PDC). PDP consists of a Mg$\^$+2/ -dependent and Ca$\^$+2)-stimulated catalytic subunit (PDPc) of Mr 52,600 and a FAD-containing regulatory subunit (PDPr) of Mr 95.600. Catalytic subunit of pyruvate dehydrogenase phosphatase (PDPc) has been suggested to have three major functional domains such as dihydrolipoamide acetyltransferase(E$_2$)-binding domain, regulatory subunit of PDP(PDPr)-binding domain, and calcium-binding domain. In order to identify functional domains, recombinant catalytic subunit of pyruvate dehydrogenase phosphatase (rPDPc) was expressed in E. coli JM101 and purified to near homogeneity using the unique property of PDPc: PDPm binds to the inner lipoyl domain (L$_2$) of E$_2$ of pyruvate dehydrogenase complex (PDC) in the presence of Ca$\^$+2/, not under EGTA. PDPc was limited-proteolysed by trypsin, chymotrypsin, Arg-C, and elastase at pH7.0 and 30$^{\circ}C$ and N-terminal analysis of the fragment was done. Chymotrypsin, trypsin, and elastase made two major framents: N-terminal large fragment, approx. 50kD and C-terminal small fragment, approx. 0 kDa. Arg-C made three major fragments: N-terminal fragment, approx. 35 kD, and central fragment, approx. 15 kD, and C-terminal fragment, approx. 10 kD. This study strongly suggest that PDPc consists of three major functional domains. However, further study should be necessary to identify the functional role.

• Bulletin of the Korean Chemical Society
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• v.28 no.5
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• pp.832-834
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• 2007

• BMB Reports
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• v.31 no.3
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• pp.303-306
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• 1998

Serine proteinase cDNA fragment from protozoan parasite Acanthamoeba culbertsoni was amplified by the reverse transcription-polymerase chain reaction (RTPCR) using degenerate oligonucleotide primers derived from conserved serine proteinase sequences. The amplified DNA fragment was subcloned and sequenced. The sequence analysis and alignment showed significant sequence similarity to other eukaryotic serine proteinases and conservation of the His, Asp, and Ser residues that form the catalytic triad. The cDNA fragment was cloned into the pGEMEX-1 expression vector and expressed in Escherichia coli. A resulting fusion protein of 56 kDa had proteolytic activity. The fusion protein reacted with sera of mice immunized with purified serine proteinase of A. culbertsoni in Western blot. Immune recognition of the fusion protein by mouse antisera suggested that the fusion protein may be valuable as a diagnostic reagent.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2001.06a
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• pp.118-119
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• 2001

A Southern-hybridization analysis and size-selected DNA library screening led to the isolation of a 6.3-kbp S. setonii DNA fragment, from which the Cl20-encoding genetic locus was found to be located within a 1.4-kbp DNA fragment. A complete nucleotide sequencing analysis of the 1.4-kbp DNA fragment revealed a 0.84-kbp ORF, which showed a strong overall amino acid similarity to the known high－G＋C gram-positive bacterial mesophilic C120s. The heterologous expression of the cloned 1.4-kbp DNA fragment in E. coli demonstrated that this Cl20 possessed a thermophilic activity within a broad temperature range and showed a higher activity against 3-methy1catechol than catechol or 4-methy-catechol, but no activity against protocatecuate.

• Journal of Microbiology
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• v.40 no.2
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• pp.146-150
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• 2002

A multiplex polymerase chain reaction (PCR) assay was developed for the identification of two Candida species-albicans and dubliniensis. Three sets of primers were selected from different genomic sequences to specifically amplify a 516 bp fragment within the tops gene, specific for several species of the genus Candida (CCL primers); a 239 bp fragment within the $\alpha$INT1 gene, specific for Candida albicans (CAL primers); and a 175 bp fragment within the ALSD1 gene, specific for Candida dubliniensis (CDL primers). Using the primers in conjunction (multiplex PCR), we were able to detect both C. albicans and C. dubliniensis and to differentiate between them. The detection limit of the PCR assay was approximately 10 cells per milliliter of saline. Thus, this multiplex PCR assay can be applied for differentiation of C. albicans and C. dubliniensis from clinical specimens.

• Environmental Mutagens and Carcinogens
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• v.18 no.2
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• pp.83-88
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• 1998

To detect the mutation in a given sequence, there are variety of methods developed by use of the gel electrophoresis. One of the methods, TGGE (Temperature Gradient Gel Electrophoresis), is a popular technique because it can detect mutations in DNA fragment with ease and at low cost. This study used 200 bp BamHI-digested DNA fragment containing the human $\varepsilon$-globin promoter which was mutated[$\varepsilon$ F1*(-141), GATA- I*(-163), and GATA-1* & $\varepsilon$F1]. This BamHI-digested DNA fragment was directly used to detect the mutated DNA fragment on 50% denaturant gel with temperature gradient of 45$^{\circ}C$ through $53^{\circ}C$. In agreement with the theoretical result of MELTSCAN program (Brossette and Wallet, 1994) the mobilities of mutated DNA fragments were shown to be nearly distinguished on the temperature gradient gel. In contrast to the above result the heteroduplex analysis under the temperature gradient condition was shown to detect the mutated DNA fragments through the heteroduplex formation between strands of mutated DNA and wild-type DNA.

• KSBB Journal
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• v.10 no.3
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• pp.264-270
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• 1995

Trigonopsis variabilis is a strong producer of D-amino acid oxidase that can transform cephalosporin C(ceph C) to ${\alpha}$-keto-adipyl-7-aminocephalosporanic acid(AKA-7ACA). Polymerase chain reaction (PCR) was applied to isolate the D-AAO gene from T. variabilis. To clone the PCR fragment, four different methods were examined using enzymatic reactions of Taq DNA polymerase, Klenow, T4 DNA polymerase I, Alkaline phosphatase Calf Intestinal, and T4 kinase. Ligation of phosphorylated blunt-end PCR fragment and dephosphorylated blunt-end of pUC18 plasmid yielded the best cloning efficiency One of recombinant E. coli transformants showed D-AAO activity against ceph C in both cell extracts and permeabilized cells.