• The Korean Journal of Malacology
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• v.31 no.4
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• pp.257-262
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• 2015

The seven selected primers BION-13, BION-29, BION-61, BION-64, BION-68, BION-72 and BION-80 generated the total number of loci, average number of loci per lane and specific loci in Meretrix lusoria (ML), Saxidomus purpuratus (SP) and Cyclina sinensis (CS) species. Here, the complexity of the banding patterns varied dramatically between the primers from the three venerid clam species. The higher fragment sizes (> 1,000 bp) are much more observed in the SP species. The primer BION-68 generated 21 unique loci to each species, which were ascertaining each species, approximately 150 bp, 300 bp and 450 bp, in the ML species. Remarkably, the primer BION-80 detected 7 shared loci by the three clam species, major and/or minor fragments of sizes 500 bp, which were matching in all samples. As regards average bandsharing value (BS) results, individuals from CS clam species (0.754) exhibited higher bandsharing values than did individuals from SP clam species (0.607) (P < 0.05). In this study, the dendrogram obtained by the seven oligonucleotides primers indicates three genetic clusters: cluster 1 (LUSORIA01-LUSORIA07), cluster 2 (PURPURATUS08-PURPURATUS14), cluster 3 (SINENSIS15-SINENSIS21). Among the twenty one venerid clams, the shortest genetic distance that displayed significant molecular differences was between individuals 18 and 20 from the CS species (genetic distance = 0.071), while the longest genetic distance among the twenty-one individuals that displayed significant molecular differences was between individuals LUSORIA no. 02 and PURPURATUS no. 09 (genetic distance = 0.778). Relatively, individuals of SP venerid species were appropriately closely related to that of CS species, as shown in the hierarchical dendrogram of genetic distances. Eventually, PCR fragments exposed in the present study may be worthwhile as a DNA marker the three venerid clam species to discriminate.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.6
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• pp.871-884
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• 2009

Gene expression profiling is a useful tool for identifying critical genes and pathways in metabolism. The objective of this study was to determine the major differences in the expression of genes associated with metabolism and metabolic regulation in liver and mammary tissues of lactating cows. We used the Michigan State University bovine metabolism (BMET) microarray; previously, we have designed a bovine metabolism-focused microarray containing known genes of metabolic interest using publicly available genomic internet database resources. This is a high-density array of 70mer oligonucleotides representing 2,349 bovine genes. The expression of 922 genes was different at p<0.05, and 398 genes (17%) were differentially expressed by two-fold or more with 222 higher in liver and 176 higher in mammary tissue. Gene ontology categories with a high percentage of genes more highly expressed in liver than mammary tissues included carbohydrate metabolism (glycolysis, glucoenogenesis, propanoate metabolism, butanoate metabolism, electron carrier and donor activity), lipid metabolism (fatty acid oxidation, chylomicron/lipid transport, bile acid metabolism, cholesterol metabolism, steroid metabolism, ketone body formation), and amino acid/nitrogen metabolism (amino acid biosynthetic process, amino acid catabolic process, urea cycle, and glutathione metabolic process). Categories with more genes highly expressed in mammary than liver tissue included amino acid and sugar transporters and MAPK, Wnt, and JAK-STAT signaling pathways. Real-time PCR analysis showed consistent results with those of microarray analysis for all 12 genes tested. In conclusion, microarray analyses clearly identified differential gene expression profiles between hepatic and mammary tissues that are consistent with the differences in metabolism of these two tissues. This study enables understanding of the molecular basis of metabolic adaptation of the liver and mammary gland during lactation in bovine species.

• Molecules and Cells
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• v.27 no.1
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• pp.47-54
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• 2009

A cDNA clone for a transcript preferentially expressed during an early phase of flooding was isolated from Nicotiana tabacum. Nucleotide sequencing of the cDNA clone identified an open reading frame that has high homology to the previously reported glycine-rich RNA-binding proteins. The open reading frame consists of 157 amino acids with an N-terminal RNA-recognition motif and a C-terminal glycine-rich domain, and thus the cDNA clone was designated as Nicotiana tabaccum glycine-rich RNA-binding protein-1 (NtGRP1). Expression of NtGRP1 was upregulated under flooding stress and also increased, but at much lower levels, under conditions of cold, drought, heat, high salt content, and abscisic acid treatment. RNA homopolymer-binding assay showed that NtGRP1 binds to all the RNA homopolymers tested with a higher affinity to poly r(G) and poly r(A) than to poly r(U) and poly r(C). Nucleic acid-binding assays showed that NtGRP1 binds to ssDNA, dsDNA, and mRNA. NtGRP1 suppressed expression of the fire luciferase gene in vitro, and the suppression of luciferase gene expression could be rescued by addition of oligonucleotides. Collectively, the data suggest NtGRP1 as a negative modulator of gene expression by binding to DNA or RNA in bulk that could be advantageous for plants in a stress condition like flooding.

• Parasites, Hosts and Diseases
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• v.31 no.3
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• pp.285-292
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• 1993

C. sinensis total RMh was containing large amount of 185 rRNA but little 285 rRNA. The size of the double-stranded cDNA synthesized from poly $(A)^{+}$ mRNA was 0.4-4.2 kb long with tapering unto 9.5 kb. Degenerated oligonucleotides (as 2 sense and 3 antisense Primers) were designed on the conserved regions of the known tropomyosin amino acid sequences. From one out of the PCR amplifications using total CDNA and matrix of primers, a specific gene product, 580 bp in size, was produced. Upon Southern hybridization of the PCR products with Schistosomn mnnsoni tropomyosin (SMTM) CDNA, only one signal appeared at the band of 580 bp product. This 580 bp product was considered to encode C. sinensis tropomyosin (CSTM) and cloned in pGEM-3Zf(-) for DNA sequencing. CSTM cDNA was 575 bp containing one open reading frame of 191 predicted amino acids, which revealed 86.3% homology with SMTM and 51.1% with rrichostronsylur coeubnlormis tropomyosin. CSTM cDNA obtained will serve as a probe in the studies of molecular cloning of CSTM.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.267-267
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• 1994

In order to obtain insight into the mechanism by which DNA containing a thymine photo-dimer is recognized by the excision repair enzyme, T$_4$ endonuclease V, we have taken NMR study of this protein and its complex with oligonucleotides. The conformations of five different DNA duplexes DNA I : d(GCGGATGGCG).d(CGCCTACCGC), DNA II d(GCGGTTGGCG) .d(CGCCAACCGC), DNA III : d(GCGGT ^ TGGCG) .d(CGCCAACCGC), DNA IV d(GCGGGCGGCG).d(CGCCCGCCGC) and DNA V d(GCGGCCGGCG) . d(CGCCGGCCGC) were studied by $^1$H NMR. The NMR spectra of these five DNA duplexes in the absence of the enzyme clearly show that the formation of a thymine dimer within the DNA induces only a minor distortion in the structure, and that the overall structure of B type DNA is retained. The photo-dimer formation is found to cause a large change in chemical shifts at the GC7 base pair, which is located at the 3'-side of the thymine dimer, accompanied by the major conformational change at the thymine dimer site. The binding of a mutant T$_4$ endonuclease V (E23Q), which is unable to digest DNA containing a thymine dimer, to the DNA duplex d(GCGGT ^ TGGCG)ㆍd(CGCCAACCGC) causes a large down-field shift in the imino proton resonance of GC7. Therefore, this position is thought to be either the crucial point of the interaction wi th T$_4$ endonuclease V, or the si to of a conformational change in the DNA caused by the binding of T$_4$ endonuclease V. Usually, it is very difficult to assign NMR peaks in DNA * protein complex because of severe peak overlaps. In order to overcome these peak overlaps, we used a method of deuterium incorporation.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.04a
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• pp.180-180
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• 1996

Previous studies have shown that kainic acid (KA) causes an elevation of hippocampal proenkephalin mRNA level. However, the role of proto-oncogene products, such as c-Fos, c-Jun and Fra proteins in the regulation of KA-induced proenkephalin mRNA increase in the hippocampus has not been well characterized. Thus, in the present study, the effect of cycloheximide (CHX) on KA-induced proenkephalin mRNA and immediate early gene products induction was examined. After pretreating with either vehicle or CHX (20 mg/kg, s.c.) for 30 min, KA (10 mg/kg) was administered s.c. The animals were sacrificed 1,2, or 8 hrs after KA administration. Total RNA and were isolated for Northern blot assay, and proteins were isolated for Western and electrophoretic gel-shift assays. First, we found that CHX inhibited KA-induced proenkephalin mRNA increase without altering intracellular proenkephalin protein level. Secondly, Western blot assays showed that KA increased c-Fos, c-Jun and Fra proteins at 1,2, and 8 hrs and CHX inhibited these immediate early gene products. Finally, electrophoretic gel shift assays revealed that KA increased both AP-1 and ENKCRE-2 DNA binding activities. Furthermore, CHX attenuated KA-induced AP-1 and ENKCRE-2 DNA binding activities. Both AP-1 and ENKCRE-2 DNA binding activities were abolished by cold AP-1 or ENKCRE-2 oligonucleotides, and further reduced by antibodies against c-Fos or c-Jun. Antibody against CREB reduced ENKCRE-2, but not AP-1, DNA binding activity. Our results suggest that on-going protein synthesis is required for elevation of hippocampal proenkephalin mRNA level induced by KA. All c-Fos, c-Jun, and Fra proteins appears to be involved in the regulation of hippocampal proenkephalin mRNA level induced by KA (This study was supported by a grant from KOSEF).

• Rapid Communication in Photoscience
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• v.3 no.4
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• pp.81-84
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• 2014

To examine the microenvironmental effect of DNA on the photosensitized reaction, the electron-donor-connecting porphyrin, meso-(9-phenanthryl)-tris(N-methyl-p-pyridinio) porphyrin (Phen-TMPyP), was synthesized. Phen-TMPyP can bind to oligonucleotides with two binding modes, depending on the DNA concentration. The fluorescence lifetime measurement of Phen-TMPyP shows a shorter component than that of the reference porphyrin without the phenanthryl moiety. However, the observed value is much longer than those of previously reported similar types of electron-donor-connecting porphyrins, suggesting that electron-transfer quenching by the phenanthryl moiety is not sufficient. The fluorescence quantum yield of Phen-TMPyP ($5{\mu}M$) decreased with an increase in DNA concentration of up to $5{\mu}M$ base pair (bp), possibly due to self-quenching through an aggregation along the DNA strand, increased with an increase in DNA concentration of more than $5{\mu}M$ bp and reached a plateau. The fluorescence quantum yield of Phen-TMPyP with a sufficient concentration of DNA was larger than that of the reference porphyrin. The singlet oxygen ($^1O_2$) generating activity of Phen-TMPyP was confirmed by the near-infrared emission spectrum measurement. The quantum yield of $^1O_2$ generation was decreased by a relatively small concentration of DNA, possibly due to the aggregation of Phen-TMPyP, and recovered with a sufficient concentration of DNA. The recovered quantum yield was rather smaller than that without DNA, indicating the quenching of $^1O_2$ by DNA. These results show that a DNA strand can stabilize the photoexcited state of a photosensitizer and, in a certain case, suppresses the $^1O_2$ generation.

• International Journal of Industrial Entomology and Biomaterials
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• v.10 no.2
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• pp.131-136
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• 2005

Beauveria bassiana F-101, which has high toxicity toward Acantholyda parki as well as Thecodiplosis japonensis, was an isolate to develop an alternative control system against the major forest pests. Up to now, in B. bassiana, only one pr1 gene has been isolated and characterized. Therefore, we here reported the identification of a pr1-like gene, which would be a factor of toxicity from B. bassiana F-101. The oligonucleotides for the amplification of the pr1-like gene, were chosen based on the conserved regions of the subtilisin family enzymes, pr1 genes of B. bassiana and Metarhizium anisopliae, and proteinase K of Tritirachium album. The cloned PCR fragment had 1111 bp including 52 bp intron. The deduced Pr1-like peptide showed a low identity with Pr1s of entomopathogenic fungi such as B. bassiana Pr1 (BbPr1) and M. anisopliae Pr1 (MaPr1) as well as the proteinase K of T. album (TaPrK). Instead, the deduced peptide had a substantially high amino acid sequence identity $(>65\%)$ with the serine proteases of Magnaporthe grisea (MgSPM1) and Podospora anserina (PaPspA). These results, therefore, appear to suggest that the putative Pr1-like peptide of B. bassiana F-101 belongs to the subtilisin-like serine protease family and may be a novel gene.

• Microbiology and Biotechnology Letters
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• v.18 no.6
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• pp.640-646
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• 1990

We have cloned the cdd gene from E. coli C600 using (cdd-) as a host. From the sequenced promoter region of E=, coli cdd gene which has been determined by Valentin-Hansen P. (1985), we synthesized the 23 mer oligonucleotides corresponding to the transcription initiation region and used as a probe for cloning of the cdd gene by Southern blotting. The isolated fragments in the blotting were introduced to the colony hybridization after transforming it into the E. coli JF611 (cdd-, pyr double mutant), and we identified the hybridized band at 27 kb long. From the original insert of 27 kb fragment in theBamHI site of pBR322, the 5.3 kb fragment containing the cdd gene was isolated by subsequent deletion and subeloning. From the derived plasmid pTK509, further deletion and subcloning were performed and clarified that the cdd gene was located in the 2.1 kb of SaZI/DraI segment in the insert of pTK605. The polypeptide encoded by the cloned DNA was appeared to be a molecular mass of 33,000.

• The Korean Journal of Malacology
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• v.24 no.1
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• pp.11-24
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• 2008

GDNA was isolated from the jedo venus clam (Protothaca jedoensis, Lischke) from Boryeong (jedo venus clam from Boryeong JVCB) and Wonsan (jedo venus clam from Wonsan; JVCW) located in the West Sea and the East Sea of Korean Peninsula, respectively and we performed clustering analyses, DNA polymorphisms and the populations genetic variations. In the present study, the seven decamer primer generated the one hundred and eleven major/minor specific bands in JVCB population and ninety four-specific bands in JVCW population. Seven primers generated the unique shared bands to each population of one hundred and seventy-six, on average of 25,1, in JVCB population from Boryeong and three hundred thirty, on average of 47,1, in JVCW population from Wonsan, respectively. The dendrogram obtained by the seven oligonucleotides primers, indicates two genetic clusters. Especially, two Protothaca between the individual WONSAN no. 12 and BORYEONG no. 10 showed the longest genetic distance (0.537) in comparison with other individuals used. Accordingly, RAPD analysis showed that the JVCB was a little more genetically diverse than the JVCW population. This result implies the genetic similarity owing to rearing in the same and/or similar circumstances or inbreeding within the JVCW population. So to speak, JVCB population may have high levels of genomic DNA variability owing to the introduction of the wild individuals from the other sites to sampling sites although it may be the geographically diverse distribution of this species. However, it was confirmed that it did not appear like that really in this study. We feel convinced that RAPD analysis discovered a significant genetic distance between two Protothaca population pairs (P<0.001). The existence of population discrimination and genetic diversity between two Protothaca populations was identified by RAPD analysis.