누에에서 새로운 항세균성 펩타이드 유전자를 탐색하기 위하여 E. coli K12로 체강 주사한 누에 유충의 cDNA 유전자 은행에서 차별화 선별로, 잠재 항세균성 펩타이드 유전자로 추정되는 BmInc8 클론을 분리하였다. BmInc8은 564bp의 크기를 가지며, 59개 아미노산을 coding하는 open reading frame과 2개의 잠정 폴리아데닐화 부위를 보유하고 있었다. BmInc8은 M. sexta에서 분리, 보고된 bactericidine 유전자와 61.2%의 DNA 상동성을 나타내는 것으로, 그 연역된 펩타이드 구조는 항세균성 펩타이드의 일종인 cecropin과 유사한 2가닥의 -helix가 Lysine-Proline 경첩부위에 의해 포개져 있는 형태를 갖는 것으로 추정되었다. 또한 cDNA 삽입 부위의 기능성 검정을 위해 원핵 발현벡터인 pT7-5를 이용하여 E. coli BL21(DE3) 균주에 형질전환하고 IPTG로 induction한 결과 E. coli BL21(DE3) 균주의 성장이 정지됨을 관찰할 수 있었다. 이상의 결과로 BmInc8은 DNA 상동성 비교, 연역 아미노산의 구조 추정 및 cDNA 삽입 부위를 이용한 transient expression 결과 항세균성 펩타이드를 coding하는 유전자임을 추정할 수 있었다. 또한 Bminc8의 cDNA 유전자 정보를 GenBank에 등록하였으며 등록 번호는 U30289이었다.

The plant-specific NAC (NAM, ATAF, and CUC)-domain proteins play important roles in plant development and stress responses. Comparative time-course expression analyses were carried out to analyze the expression levels of 62 soybean NAC genes during drought stress in order to search for the stress-inducible NAC genes. Ten GmSNAC (Glycine max stress-inducible NAC) genes having the significant differential expression in response to the drought stress and abscisic acid (ABA) hormone application were further investigated for their expression profiles with various stresses such as drought, high salinity, cold and with ABA treatments by the quantitative real-time PCR analyses. In this research, the full-length cDNAs of eight GmSNAC were isolated for the further studies. Eight GmSNAC proteins were tested for their transcription activation in the yeast assay system. Two GmSNAC proteins showed the very high transcriptional activities and the other two GmSNAC proteins displayed moderate levels of transactivation while the remaining four GmSNAC proteins lacked transactivation in yeast. Subcellular localization of eight GmSNAC proteins was analyzed via the green fluorescent protein-GmSNAC fusion protein in tobacco plant cell. Three GmSNAC proteins with the C-terminal transmembrane domain were localized to the nucleus and cytoplasmic fractions. The other five GmSNAC proteins were targeted to the nucleus. The function of GmSNAC49 gene was further investigated using the overexpression transgenic Arabidopsis. Germination rate in transgenic plants over-expressing GmSNAC49 was delayed in the media supplemented with mannitol or ABA compared with that of wild-type (WT) plants. The 35S:GmSNAC49 transgenic Arabidopsis displayed improved tolerance to drought stress compared to the WT. The results of this systematic analysis of the GmSNAC family responsive to abiotic stress will provide novel tools and resources for the development of improved drought tolerant transgenic soybean cultivars

The inflammatory response to infections, such as bacteria and viruses is mediated by multiple host factors. The tumor related-genes are the important cytokines in mammals. However, a number of tumor related-genes are not identified in the rock bream. Here, we have reported the identification and molecular characterization of the tumor related genes. The LPS-induced TNF-α factor 1 and 2 (LITAF1, LITAF2), tumor necrosis factor superfamily member 14 (TNFSF14), tumor necrosis factor receptor superfamily member 14 (TNFRSF14) and translationally controlled tumor protein (TCTP), programmed cell death 10 (PCD10) from rock bream are used for the under investigations. The LITAF1 and LITAF2 consist of 138 and 163 amino acids with a conserved LITAF domain. TNFSF14 and TNFRSF14 comprise 266 and 181 amino acid, respectively. TCTP encompasses of 170 amino acid containing two conserved TCTP signatures. Furthermore PCD10 consists of 210 amino acids. Using quantitative real-time PCR, we have obtained expression analysis results of LITAF1 and 2, TNFSF14, TNFRSF14, TCTP, PCD10 in the various tissue. Compared to the control, the tumor related genes mRNA is detected at a high levels in gill (LITAF1, TCTP), intestine (LITAF2), liver (PCD10), spleen (TNFSF14) and RBC (TNFRSF14). We have also performed gene expression analysis in the kidney, spleen, liver and gill after challenging with Streptococcus iniae, Edwardsiella tarda and Red seabream iridovirus. We have acquired the dynamic regulated mRNA expression to each of pathogen according to the tissue. Expression of tumor related-genes mRNA are significantly increased by infected with pathogens in most of the tissue. But oddly, PCD10 mRNA is expressed significantly decreased by S. iniae infection in all of tissues. Our results reveal that rock bream tumor relatived-genes may be involved in rock bream immune responses to pathogen infections, as well as, they also act like potential biomarkers for innate immunity.

To investigate the involvement of prepro-proopiomelanocortins (POMCs) with hypermelanosi on the blind side of olive flounder, Paralichthys olivaceus, we surveyed tissue sites in which the three POMCs genes are expressed, and compared pituitary-ofPOMCs (ofPOMC-Ⅰ, Ⅱ and Ⅲ) mRNA activities between the normal and the hypermelano flounders. For the studies, we refereed to the information of ofPOMC-Ⅰ and Ⅱ genes in NCBI, but we try to newly discover ofPOMC-Ⅲ sequence. At first, we cloned and characterized cDNA encoding its ofPOMC-Ⅲ from the pituitary of olive flounder The ofPOMC-Ⅲ cDNA consists of 1,258 bp nucleotides and 7 peptides (Signal peptide, N-POMC, α-MSH, CLIP, N-β-LPH, β-MSH and β-END-like peptide) encoded in 231 amino acids. In the test of tissue distribution of ofPOMCs in olive flounder, we found that ofPOMC-Ⅰ is expressed in pitutary only, but ofPOMC-Ⅱ and Ⅲ are expressed in all tissues examined. In the comparison of ofPOMCs mRNA expression between the normal and the hypermelano fish, expression of ofPOMC-Ⅰ, Ⅱ mRNA was significantly higher in the hypermelano group than in normal group, but ofPOMC-Ⅲ was not significantly different between two groups.

Translationally controlled tumor protein (TCTP) is one of important immune regulator. TCTP has been implicated in cellular processes including the cell growth, cell cycle progression, apoptosis regulation and the protection of cells against various stress condition. In this study, we cloned and characterized TCTP from rock bream (Oplegnathus fasciatus), which is an economically important species in the Korea aquaculture industry. The Full-length of rock bream TCTP (RbTCTP) cDNA was 1041 bp and contained an open reading frame (ORF) of 513 bp, which encoded 170 amino acid sequence. The 5' untranslated region (UTR) was 90 bp while the 3' UTR was 538 bp, containing a polyadenylation signal (ATTAAA). The identity of amino acid sequence was 76%, 75% and 74% in tilapia, orange-spotted grouper and Japanese seaperch, respectively. The positions of microtuble-binding region, Ca⁺ binding region and TCTP signature regions in RbTCTP were similar with those of other fish species and mammalian. The RbTCTP mRNA was expressed highest in the muscle. Expression of TCTP mRNA were significantly variable according to injection of red seabream iridovirus (RSIV), Streptococcosis (S. iniae) and Edwardsiella tarda (E. tarda).

미성숙 종자로부터 추출된 전체 RNA를 이용하여 합성한 cDNA와 LMW-GS 특이 프라이머세트를 이용하여 43개의 LMW-GS 유전자를 분리하였다. 각각의 유추 아미노산은 상동성이 높은 20개의 시그널 펩타이드, N-말단 영역, 반복서열영역 그리고 C-말단 영역을 가지며 C-말단 영역에 분자내 혹은 분자간 이황화 결합을 형성하는 전형적인 8개의 시스테인을 가지고 있었다. 이들 시스테인의 위치는 첫번째, 일곱번째를 제외하고는 보존되어 있었다. Ikeda

The transcription factors, DMRT1 and FOXL2, play a role in fish sex differentiation of the bipotential precursor into the male and female pathway, respectively. In order to provide the molecular background for understanding hormonal regulation in sexual determination and differentiation in the red-spotted grouper, Epinephelus akaara, one of commercially important epinephelines, and is often used to study protogynous sex change. First, we amplified the partial sequence of two genes (DMRT1 and FOXL2) from the gonad of red-spotted grouper. Also, we surveyed the tissue-specific and sex-specific expression pattern of each genes by RT-PCR. The effects of 17α-methyltestosterone (MT) in the sexually immature gonad of red-spotted grouper were investigated by Real-time quantitative RT-PCR. Fish were treated with MT-dipping method from around 70 days after hatching (DAH) for two month. DMRT1 and FOXL2 cDNA flagments consist of 489 and 836 base pairs (bp) and encodes a protein of 162 and 278 amino acids, respectively. RT-PCR revealed that DMRT1 mRNA was expressed higher level in the testis. Foxl2 was expressed extensively in the neural and peripheral tissues with its highest level in the ovary, indicating a potential role for Foxl2 in the brain-pituitary-gonad axis. Real-time quantitative RT-PCR analyses showed that DMRT1 mRNA expression was upregulated in the MT-treated fish. These results suggest that the sex inversion of red-spotted grouper by MT might be due to the suppression of FOXL2 gene expression, and resulting in the induction of the 11-KT secretion.

Cinnamyl alcohol dehydrogenase (CAD, EC 1.1.1.95), catalyzes the reduction of hydroxycinnamaldehydes to give hydroxycinnamyl alcohols, or "monolignols," the monomeric precursors of lignin. Lignins are important components of cell walls and lignified secondary cell walls play crucial roles in long distance transport of water and nutrients during plant growth and development and in plant defense against biotic and abiotic stresses. Here a cDNA clone containing a CAD gene, named as PgCAD, was isolated from a commercial medicinal plant Panax ginseng. PgCAD is predicted to encode a precursor protein of 177 amino acid residues, and its sequence shares high homology with a number of other plant CADS. The expression of PgCAD in adventitious roots and hairy roots of P. ginseng was analyzed using reverse transcriptase (RT)-PCR under various abiotic stresses such as salt, salicylic acid, wounding and chilling treatment that triggered a significant induction of PgCAD at different time points within 2-48 h post-treatment. This study revealed that PgCAD may help the plants to survive against various abiotic stresses.

We isolated low temperature inducible genes using suppression subtractive hybridization (SSH) method and were able to obtain to clone MLT7 gene encoding peroxiredoxin and aminotransferase. The full-length cDNA of MLT7 is 1,049 bp with an open reading frame (ORF) consisting of 261 amino acid (aa). Genomic southern blot confirmed that mungbean genome has two copies of MLT7 gene. Northern blot analysis was also carried out for the gene expression during ABA, NaCl, drought, wounding and H2O2 stresses. The expression of MLT7 gene significantly decreased by ABA, NaCl and drought stress, but wounding and H2O2 stress significantly induced MLT7 gene expression. Especially, H2O2 strongly induced the MLT7 gene expression. The expression of MLT7 gene during low temperature stress started to increase in 3 h after treatment, and than slightly decreased and again increased at 24 h. Using GFP fusion vector, GFP-MLT7 was targeted both to mitochondria and chloroplast. However, it was mostly targeted to mitochondria and partially targeted to chloroplast. For the functional analysis of MLT7, MLT7 recombinant protein was heterologously expressed in E. coli. The MLT7 recombinant cells showed enhanced antioxidant activity compared to that of vector control cells.