• Journal of Animal Science and Technology
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• v.45 no.6
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• pp.933-938
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• 2003

This study was performed to analyse the sequences of variations of mtDNA D-loop and their effects on carcass traits in Hnawoo(Korean cattle). The resulting sequences were compared with previously published sequences for other cattle breeds(GenBank J01394). The PCR was used to amplify a total of 964 bp between nucleotide 15758 and 383 within D-loop region of mtDNA using specific primers. Twenty five polymorphic sites by nucleotide substitution were found in mtDNA of Hanwoo. The frequencies of positions at 169, 16042, 16093, 16119, 16255 and 16302 nt with high levels of sequence polymorphism were 0.891, 0.117, 0.109, 0.182, 0.197 and 0.117, respectively. The substitution effect at 169 and 16119 nt was found significant on marbling score. Also substitution effect at 169 and 16042 nt was highly significant(p〈0.01) on backfat. thickness. Polymorphism of mtDNA sequence in D-loop region could be useful for the analysis of cytoplasmic genetic variation and associations with the other economically important traits and maternal lineage analysis in Hanwoo.

• Korean Journal of Microbiology
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• v.41 no.2
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• pp.140-145
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• 2005

Streptomyces natalensis ATCC27448 produces natamycin, a commercially important macrolide antifungal antibiotic. For molecular genetic study of S. natalensis, we have developed a system for introducing DNA into S. natalensis via conjugal transfer from Escherichia coli. An effective transformation procedure for S. natalensis was established based on transconjugation from E, coli ET12567/pUZ8002 using a ${\Phi}C31$-derived integration vector, pSET152, containing oriT and attP fragments. The high frequency was obtained on MS medium containing 10 mM $MgCl_2$ using $6.25\times10^8$ of E.coli donor cells without heat treatment of spores. In addition, southern blot analysis of exconjugants and the sequence of plasmids containing DNA flanking the insertion sites from the chromosome revealed that S. natalensis contains a single ${\Phi}C31$ attB site and at least a secondary or pseudo attB site. Similar to the case of various Streptomyces species, a single ${\Phi}C31$ attB site of S. natalensis is present within an ORF encoding a pirin-homolog, but a pseudo-attB site is present within a distinct site (GenBank accession no. $YP\_117731$) and also its sequence deviates from the consensus sequences of attB sequence.

• Journal of Microbiology and Biotechnology
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• v.14 no.3
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• pp.620-627
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• 2004

On the foundation of a database of genome sequences and protein analyses, the ability to clone a gene based on a peptide analysis is becoming more feasible and effective for identifying a specific gene and its protein product of interest. As such, the current study conducted a protein analysis using 2-D PAGE followed by MALDI- TOF and ESI-MS to identify a highly expressed gene product of C. parasitica. A distinctive and highly expressed protein spot with a molecular size of 47.2 kDa was randomly selected and MALDI-TOF MS analysis was conducted. A homology search indicated that the protein appeared to be a fungal enolase (enol). Meanwhile, multiple alignments of fungal enolases revealed a conserved amino acid sequence, from which degenerated primers were designed. A screening of the genomic $\lambda$ library of C. parasitica, using the PCR amplicon as a probe, was conducted to obtain the full-length gene, while RT-PCR was performed for the cDNA. The E. coli-expressed eno 1 exhibited enolase enzymatic activity, indicating that the cloned gene encoded the C. parasitica enolase. Moreover, ESI-MS of two of the separated peptides resolved from the protein spot on 2-D PAGE revealed sequences identical to the deduced sequences, suggesting that the cloned gene indeed encoded the resolved protein spot. Northern blot analysis indicated a consistent accumulation of an eno1 transcript during the cultivation.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.5
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• pp.638-644
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• 2007

Differences of meat qualities between Japanese Black and Holstein have been known in Japan, however, the causative proteins and/or the genetic background have been unclear. The aim of this study was to identify candidate proteins causing differences of the meat qualities between the two breeds. Using technique of two-dimensional gel electrophoresis, protein profiling was compared from samples of the longissimus dorsi muscle and subcutaneous adipose tissue. Five protein spots were observed with different expression levels between breeds. By using LC-MS/MS analysis and Mascot program, three of them were identified as ankyrin repeat protein 2, phosphoylated myosin light chain 2 and mimecan protein. Subsequently, we compared the DNA coding sequences of three proteins between breeds to find any nucleotide substitution. However, there was no notable mutation which could affect pI or molecular mass of the proteins. The identified proteins may be responsible for different characteristics of the meat qualities between Japanese Black and Holstein cattle.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.2
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• pp.581-590
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• 2016

Resistance to anoikis, a cell-detachment induced apoptosis, is one of the malignant phenotypes which support tumor metastasis. Molecular mechanisms underlying the establishment of this phenotype require further investigation. This study aimed at exploring protein expression profiles associated with anoikis resistance of a metastatic breast cancer cell. Cell survival of suspension cultures of non-metastatic MCF-7 and metastatic MDA-MB-231 cells were compared with their adherent cultures. Trypan blue exclusion assays demonstrated a significantly higher percentage of viable cells in MDA-MB-231 than MCF-7 cell cultures, consistent with analysis of annexin V-7-AAD stained cells indicating that MDA-MB-231 possess anti-apoptotic ability 1.7 fold higher than MCF-7 cells. GeLC-MS/MS analysis of protein lysates of MDA-MB-231 and MCF-7 cells grown under both culture conditions identified 925 proteins which are differentially expressed, 54 of which were expressed only in suspended and adherent MDA-MB-231 but not in MCF-7 cells. These proteins have been implicated in various cellular processes, including DNA replication and repair, transcription, translation, protein modification, cytoskeleton, transport and cell signaling. Analysis based on the STITCH database predicted the interaction of phospholipases, PLC and PLD, and 14-3-3 beta/alpha, YWHAB, with the intrinsic and extrinsic apoptotic signaling network, suggesting putative roles in controlling anti-anoikis ability. MDA-MB-231 cells grown in the presence of inhibitors of phospholipase C, U73122, and phospholipase D, FIPI, demonstrated reduced ability to survive in suspension culture, indicating functional roles of PLC and PLD in the process of anti-anoikis. Our study identified intracellular mediators potentially associated with establishment of anoikis resistance of metastatic cells. These proteins require further clarification as prognostic and therapeutic targets for advanced breast cancer.

• Molecules and Cells
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• v.40 no.9
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• pp.621-631
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• 2017

Vaccinia-related kinase 1 (VRK1) and VRK3 are members of the VRK family of serine/threonine kinases and are principally localized in the nucleus. Despite the crucial roles of VRK1/VRK3 in physiology and disease, the molecular and functional interactions of VRK1/VRK3 are poorly understood. Here, we identified over 200 unreported VRK1/VRK3-interacting candidate proteins by affinity purification and LC-MS/MS. The networks of VRK1 and VRK3 interactomes were found to be associated with important biological processes such as the cell cycle, DNA repair, chromatin assembly, and RNA processing. Interactions of interacting proteins with VRK1/VRK3 were confirmed by biochemical assays. We also found that phosphorylations of XRCC5 were regulated by both VRK1/VRK3, and that of CCNB1 was regulated by VRK3. In liver cancer cells and tissues, VRK1/VRK3 were highly upregulated and its depletion affected cell cycle progression in the different phases. VRK3 seemed to affect S phase progression and G2 or M phase entry and exit, whereas VRK1 affects G1/S transition in the liver cancer, which could be explained by different interacting candidate proteins. Thus, this study not only provides a resource for investigating the unidentified functions of VRK1/VRK3, but also an insight into the regulatory roles of VRK1/VRK3 in biological processes.

• Korean Journal of Agricultural Science
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• v.41 no.3
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• pp.187-191
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• 2014

The cultivation area and use of genetically modified (GM) crops have been increased continuously over the world. Concerns about the potential risks of GM crops are also increasing. Safe management for the development and production of GM crops is required according to Living Modified Organism Act in Korea. Planning about the methods, duration, and frequency of environmental monitoring is also required for commercial use of GM crops. GM Zoysia japonica Steud. (event name: JG21) expressing resistance to glufosinate-ammonium has been generated previously. By using gamma ray treatment to JG21 we also developed male sterility and dwarf Z. japonica (event name: JG21-MS). The objective of this study was to establish the monitoring system for environment release of JG21-MS. In this study we extracted RNA from JG21 and JG21-MS and conducted RAPD (random amplified polymorphic DNA) method to distinguish JG21 and JG21-MS.

• Korean Journal of Plant Tissue Culture
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• v.26 no.1
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• pp.21-26
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• 1999

These experiments were attempted to introduce rolC gene in the Petunia hybrida cv. Titan white by Agrobacterium mediated. The maximum frequency of shoot regeneration was obtained by 60% on MS medium containing 1.0 mg/L BA, 0.1 mg/L NAA, 200 mg/L kanamycin, 500 mg/L carbenicillin, 30 g/L sucrose, and 8 g/L agar. Kanamycin-resistant calli were selected from petunia leaf discs by cocultivation with Agrobacterium suspension cultures on MS medium. The addition of AgNO$_3$ and KMnO$_4$ in the medium increased the shoot regeneration by 31.3% from leaf disc as compared with non-treated leaf disc. Among clones exhibiting kanamycin resistance, only 3 clones were confirmed by southern hybridization analysis.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.3
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• pp.328-333
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• 2007

Carboxypeptidase E (CPE) plays an important role in the regulation of the body fat content. Therefore, it has been suggested as candidate gene for traits related to meat quality in beef cattle. This study was conducted to identify single nucleotide polymorphisms (SNPs) in the CPE gene and to investigate association of SNP marker with carcass and meat quality traits in Korean cattle. Three SNPs were identified in the intron 4 (A309G SNP and C445T SNP) and exon 5 (C601T SNP) of the CPE gene by sequence analyses of CPE cDNA and genomic DNA samples. The sequences have been deposited in GenBank database with accession numbers AY970664 and AY970663. Genotyping of the gene-specific SNP marker was carried out using the PCR-RFLP with restriction enzymes DdeI for C445T SNP and NlaIII for C601T SNP. The frequencies of C and T alleles were 0.43 and 0.57 for C445T SNP and 0.42 and 0.58 for C601T SNP, respectively. Statistical analysis indicated that the C445T SNP showed a significant effect (p<0.05) on marbling score (MS) and breeding value of backfat thickness (BF-EBV), respectively. Animals with the CT genotype showed higher marbling score and backfat thickness than those with the TT genotype. This marker also showed a significant dominance effect for the MS and BF-EBV (p<0.05). However, no significant associations were observed between C601T SNP genotypes and all traits examined. The results suggest that the CPE gene may be used as a marker for carcass traits in Korean cattle.

• Parasites, Hosts and Diseases
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• v.51 no.3
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• pp.327-334
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• 2013

Vaccination is considered a promising alternative for controlling tick infestations. Haemaphysalis longicornis midgut proteins separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membrane were screened for protective value against bites. The western blot demonstrated the immunogenicity of 92 kDa protein (P92). The analysis of the P92 amino acid sequence by LC-MS/MS indicated that it was a H. longicornis paramyosin (Hl-Pmy). The full lenghth cDNA of Hl-Pmy was obtained by rapid amplification of cDNA ends (RACE) which consisted of 2,783 bp with a 161 bp 3' untranslated region. Sequence alignment of tick paramyosin (Pmy) showed that Hl-Pmy shared a high level of conservation among ticks. Comparison with the protective epitope sequence of other invertebrate Pmy, it was calculated that the protective epitope of Hl-Pmy was a peptide (LEEAEGSSETVVEMNKKRDTE) named LEE, which was close to the N-terminal of Hl-Pmy protein. The secondary structure analysis suggested that LEE had non-helical segments within an ${\alpha}$-helical structure. These results provide the basis for developing a vaccine against biting H. longicornis ticks.