• BMB Reports
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• 제39권5호
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• pp.607-617
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• 2006

A novel calcium-dependent protein kinase gene (designated as IiCPK2) was cloned from tetraploid Isatis indigotica. The full-length cDNA of IiCPK2 was 2585 bp long with an open reading frame (ORF) of 1878 bp encoding a polypeptide of 625 amino acid residues. The predicted IiCPK2 polypeptide included three domains: a kinase domain, a junction domain (or autoinhibitory region), and a C-terminal calmodulin-like domain (or calcium-binding domain), which presented a typical structure of plant CDPKs. Further analysis of IiCPK2 genomic DNA revealed that it contained 7 exons, 6 introns and the length of most exons was highly conserved. Semi-quantitative RT-PCR revealed that the expression of IiCPK2 in root, stem and leaf were much higher in tetraploid sample than that in diploid progenitor. Further expression analysis revealed that gibberellin ($GA_3$), NaCl and cold treatments could up-regulate the IiCPK2 transcription. All our findings suggest that IiCPK2 might participate in the cold, high salinity and GA3 responsive pathways.

• 한국어병학회지
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• 제22권3호
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• pp.343-352
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• 2009

Natural-killer-cell-enhancing factor (NKEF) belongs to the newly defined peroxiredoxin (Prx) family. It was originally isolated from human erythroid cells. The black rockfish NKEF cDNA was identified through the expressed sequence tag (EST) analysis of PBLs libraries. The full-length NKEF cDNA was 1433 bp long and contained an open reading frame (ORF) of 594 bp that encoded 198 amino-acid residues. The 5' UTR had a length of 39 bp, and the 3’UTR 800 bp. The deduced amino-acid sequence of the black rockfish had a density 93.4, 92.9, 87.8, 85.8, 84.8, 83.8, 80.3, 79.7, 77.2, and 75.2% that of the pufferfish, olive flounder, channel catfish, zebrafish, chicken, common carp, Myotis lucifugus, cattle, human PrxI, rat PrxI, human NKEF-A, and Xenopus tropicalis, respectively. The NKEF gene was expressed in all the tissues of the black rockfish. The RT-PCR indicated that the NKEF transcripts were predominantly in the spleen and gill, less dominantly in the PBLs, head kidney, trunk kidney, and liver, and least in the intestine and muscles. This is the first report on the existence of the NKEF-A gene in black rockfish.

• 환경생물
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• 제27권4호
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• pp.406-413
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• 2009

The enzyme invertase contributes to sugar unloading, pathogen defense, differentiation and development in plants. We cloned the complete cDNA of a soluble acid invertase from pea seedlings (Pisum sativum) via RT-PCR and the rapid amplification of the cDNA end (RACE) technique. The full-length cDNA of the soluble pea invertase comprised 2237 bp and contained a complete open reading frame encoding 647 amino acids. The deduced amino acid sequence showed high homology to soluble acid invertases from various plants. Northern blot analysis demonstrated the soluble acid invertase gene of P. sativum was strongly expressed in sink organs such as shoot tips and root tips, and induced by abscisic acid, gibberellic acid and jasmonic acid in shoots. Especially, gibberellic acid enhanced the gene expression of the soluble acid invertase in a time-dependent manner. This study presents that the gene expression patterns of a soluble acid invertase from pea are strongly consistent with the suggestion that individual invertase gene product has different functions in the growing plant.

• Molecules and Cells
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• 제19권2호
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• pp.198-204
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• 2005

Both erythropoietin (EPO) and the short-form thrombopoietin (TPO) were expressed at low levels whereas the long-form TPO was expressed at high levels in transgenic animals. To elucidate the role of carboxy-terminal half of the long-form TPO which is absent in the short-form, we generated recombinant TPO or EPO expression vectors which contain or lack the carboxy-terminal half of TPO and examined their expression in the HC11 and 293 cells. The long-form TPO was expressed higher than the short-form regardless of the cell types, transfection modes, and promoters. When 3'-half of the long-form TPO cDNA was placed downstream of the EPO cDNA to act as a 3'-untranslated region, expression of EPO was moderately increased at the RNA level, however, no remarkable increase was observed at the protein level. These results suggest that the low expression of EPO, as like as the short-form TPO, is due to absence of the 3'-half in the full-length TPO that confers stability both at the RNA and protein levels.

• International Journal of Industrial Entomology and Biomaterials
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• 제21권1호
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• pp.127-132
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• 2010

We have characterized full-length cDNA encoding Gall-6-tox protein, which was cloned from the fat body of the immunized Galleria mellonella larvae. The cloned cDNA of Gall-6-tox consists of 1301 nucleotides and contained an open reading frame of 891 nucleotides corresponding to a protein of 296 residues that includes a putative 16-residue signal sequence and a 280-residue mature peptide with a calculated mass of 30,707.73 Da. The deduced mature peptide contains conserved tandem repeats of six cysteine-stabilized alpha beta ($Cs{\alpha}{\beta}$) motifs, which was detected in scorpion toxins and insect defensins. In the sequence homology search, mature Gall-6-tox showed 34% and 28% amino acid sequence homology with Bomb-6-tox from Bombyx mori and Spod-11-tox from Spodoptera frugiperda, respectively. Gall-6-tox orthologs were only found in Lepidopteran species, indicating that this new immune-related gene family is specific to this insect order. RT-PCR analysis revealed that Gall-6-tox was expressed primarily in the larval fat bodies, hemocytes, and midgut against invading bacteria into hemocoel. Moreover, the expression time course of Gall-6-tox was examined up to 24 h in the fat bodies and midgut after injection of E. coli. Altogether, these results suggest that Gall-6-tox is derived from defensins and Gall-6-tox may play a critical role in Lepidoptera immune system.

• Asian Pacific Journal of Cancer Prevention
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• 제15권12호
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• pp.5055-5061
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• 2014

In this study, an anti-oxidant and anti-tumor protein Latcripin-3 of Lentinula edodes C91-3 was expressed in Escherichia coli. for the first time. According to the cDNA library, the full-length gene of Latcripin-3 was cloned by the methods of 3'-full rapid amplification of cDNA Ends (RACE) and 5'-full RACE. The structural domain gene of Latcripin-3 was inserted into the pET32 a(+). The functional protein of Latcripin-3 was expressed in Rosetta-gami (DE3) E. coli, evaluated by Western blotting and mass spectrometry. DPPH testing showed that the protein Latcripin-3 can scavenge free radicals remarkably well. The activity of functional protein Latcripin-3 on A549 cells was studied with flow cytometry and the MTT method. The MTT assay results showed that there was a decreases in cell viability in a dose-dependent and time-dependent manner in protein Latcripin-3 treated groups. Flow cytometry demonstrated that Latcripin-3 can induce apoptosis and block S phase dramatically in human A549 lung cancer cells as compared to the control group. At the same time, the cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. This research offers new insights and advantages for identifying anti-oxidant and anti-tumor proteins.

• 한국어병학회지
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• 제14권2호
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• pp.59-64
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• 2001

두툽상어의 간조직 cDNA library의 EST를 통해 중금속의 세포내 농도 조절과 환경으로부터 흡수한 유해 중금속의 해독작용 등의 기능을 수행하는 MT 유전자를 cloning하였다. 염기서열 분석 결과 두툽상어의 MT 유전자는 204bp의 coding 영역과 182bp의 3'UTR 영역으로 구성되어 있었으며, 종결코돈 이후 162bp의 polyadenylatin 신호서열과 이로부터 15bp 이후의 poly(A)서열 등이 확인되었다. 염기서열로부터 유추한 68개의 아미노산 서열에는 다른 척추동물에서와 같이 Cys 잔기가 전체의 29.4%(20/68)로 풍부하였으며, 아미노산 서열수준에서 포유류와는 43~54%, 어류들과는 41~45%의 상동성을 나타냈었다. 특히 20개의 Cys은 어류와 18개가 다른 척추동물과는 19개가 잘 보존되어 있었다. 두툽상어 MT는 특이하게 모든 척추동물에서 잘 보존된 $\beta$-domain 끝 쪽의 9번째 Cys앞에 5개의 아미노산을 더 갖고 있었으며, 경골어류 MT의 특징인 4번째 위치의 gap이 없고, 18번째 Cys의 위치가 어류와 달리 다른 척추동물들과 같았다. 또한 C 말단의 아미노산 잔기가 다른 생물체와는 모두 다른 Ser을 갖는 특징을 나타내었다. 이와 같은 두툽상어 MT유전자의 특징들은 연골어류의 분자적 진화과정을 알 수 있는 분자 표지유전자로 이용할 수 있을 것이다.

• Fisheries and Aquatic Sciences
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• 제10권2호
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• pp.60-67
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• 2007

Myostatin (MSTN; also known as GDF8) is a member of the transforming growth factor ${\beta}-superfamily$ of proteins. MSTN negatively regulates mammalian skeletal muscle growth and development by inhibiting myoblast proliferation. Mice and cattle possessing mutant MSTN alleles display a 'double muscling' phenotype characterized by extreme skeletal muscle hypertrophy and/or hyperplasia. We isolated the full-length cDNA of a novel MSTN gene from S. schlegeli muscle tissue and examined its expression pattern in various tissues. The full-length gene (GenBank DQ423474) consists of 1941bp with an open reading frame of 1134 bp, encoding 377 amino acids that show 62-92% amino acid similarity to other vertebrate MSTNs. The predicted protein contains a conserved proteolytic cleavage site (RXRR) and nine conserved cysteine residues at the C terminus. RT-PCR revealed that the unprocessed and prodomain myostatin mRNAs were predominantly present in muscle, with limited expression in other tissues. However, the mature myostatin mRNA was highly expressed in brain and muscle, intermediately expressed in the gills, intestine, heart, and kidney, and weakly expressed in the liver and spleen.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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• pp.146.2-147
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• 2003

Two Korean isolates of Alfalfa mosaic virus (AHV-AZ, AMV-KR) were isolated from azuki bean and potato plants, respectively, and their pathologies were confirmed on some susceptible host plants including pepper, tobacco and red bean plants. Full length cDNAs to RNA1, RNA2 and RNA3 of the two Korean strains were amplified using the long-template reverse transcription (RT)-polymerase chain reaction (PCR) method. RT-PCR products covering entire regions for the three AMV genome RNAs were cloned. RNA transcripts were synthesized in vitro from each clones using T7 RNA polymerase and infectivity test was peformed in 9 reassortment sets of transcripts. All the combinations of reassorted transcripts were found to be infectious when inoculated onto Nicotiana benthamiana plants, and were not distinguishable to those of wild types. The full-length cDNA clones that were confirmed infectious were sequenced their nucleotide sequences. We will discuss sequence analysis of the two Korean isolates of AMV genomic RNA3 and compare reported foreign isolates of AMV.

• Journal of Microbiology and Biotechnology
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• 제17권7호
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• pp.1106-1112
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• 2007

This report provides the complete nucleotide sequences of the full-length cDNA encoding squalene synthase (SQS) and its genomic DNA sequence from a triterpene-producing fungus, Ganoderma lucidum. The cDNA of the squalene synthase (SQS) (GenBank Accession Number: DQ494674) was found to contain an open reading frame (ORF) of 1,404 bp encoding a 468-amino-acid polypeptide, whereas the SQS genomic DNA sequence (GenBank Accession Number: DQ494675) consisted of 1,984 bp and contained four exons and three introns. Only one gene copy was present in the G. lucidum genome. The deduced amino acid sequence of Ganoderma lucidum squalene synthase (GI-SQS) exhibited a high homology with other fungal squalene synthase genes and contained six conserved domains. A phylogenetic analysis revealed that G. lucidum SQS belonged to the fungi SQS group, and was more closely related to the SQS of U. maydis than to those of other fungi. A gene expression analysis showed that the expression level was relatively low in mycelia incubated for 12 days, increased after 14 to 20 days of incubation, and reached a relatively high level in the mushroom primordia. Functional complementation of GI-SQS in a SQS-deficient strain of Saccharomyces cerevisiae confirmed that the cloned cDNA encoded a squalene synthase.