Lactic acid bacteria (LAB) are widely used for various food fermentation. With the recent advances in modern biotechnology, a variety of bio-products with the high economic values have been produced using microorganisms. For molecular cloning and expression studies on the gene of interest, E. coli has been widely used mainly because vector systems are fully developed. Most plasmid vectors currently used for E, coli carry antibiotic-resistant markers. As it is generally believed that the antibiotic resistance markers are potentially transferred to other bacteria, application of the plasmid vectors carrying antibiotic resistance genes as selection markers should be avoided, especially for human consump-tion. By contrast, as LAB have some desirable traits such that the they are GRAS(generally recognized as safe), able to secrete gene products out of cell, and their low protease activities, they are regarded as an ideal organism for the genetic manipulation, including cloning and expression of homologous and heterologous genes. However, the vec-tor systems established for LAB are stil insufficient to over-produce gene products, stably, limiting the use of these organisms for industrial applications. For a past decade, the two popular plasmid vectors, pAM$\beta$1 of Streptococcus faecalis and pGK12 theB. subtilis-E. coli shuttle vector derived from pWV01 of Lactococcus lactis ssp. cremoris wg 2, were most widely used to construct efficient chimeric vectors to be stably maintained in many industrial strains of LAB. Currently, non-antibiotic markers such as nisin resistance($Nis^{r}$ ) are explored for selecting recombi-nant clone. In addition, a gene encoding S-layer protein, slp/A, on bacterial cell wall was successfully recombined with the proper LAB vectors LAB vectors for excretion of the heterologous gene product from LAB Many food-grade host vec-tor systems were successfully developed, which allowed stable integration of multiple plasmid copies in the vec-mosome of LAB. More recently, an integration vector system based on the site-specific integration apparatus of temperate lactococcal bacteriophage, containing the integrase gene(int) and phage attachment site(attP), was pub-lished. In conclusion, when various vector system, which are maintain stably and expressed strongly in LAB, are developed, lost of such food products as enzymes, pharmaceuticals, bioactive food ingredients for human consump-tion would be produced at a full scale in LAB.

Aspergil-lus coreanus NR-15 and Aspergilus oryzae NR-2-5 from traditional Korean Nuruk were selected as parental strains producing starch hydrolysis enzyme. Xll(Arginine-) mutant from A. coreanus NR 15-1 showed high glu-doamylase activity and total acid productivity. Z6(Adenine-) mutant from A. oryzae NR2-5 showed the highest $\alpha$-amylase activity. Therefore, both XII and Z6 mutants were selected and investigated for the optimal conditions of protoplast formation for protoplast fusion. Mixture of equal amount of cellulase and driselase(10mg/ml each) was the most effective as lytic enzymes. The optimal pH and temperature for protoplast formation were 5.0 and $30^{\circ}C$, respectively. The most effective reaction for protoplast formation time was 4 hours. The maximum of protoplst for- mation of Xll mutant and Z6 mutant were $6.54$\times$10^{7}$ protoplasts/ ml and $3.04$\times$10^{ 7}$ protoplasts/ml, and the regen-eration frequencies of the protoplasts were 11.3% and 11.6%, respectively. The size of the protoplasts from X11 and Z6 mutants were 3~6 $\mu\textrm{m}$ and 4~9$\mu\textrm{m}$, respectively.

누에 핵다각체병 바이러스의 gp64 프로모터를 이용한 transient 발현 벡터를 제작하기 위해서 gp64 유전자의 구조를 분석하였다. Southern blotting 분석을 통해 genome DNA에서 gp64 유전자를 탐색하기 gp64 구조유전자를 포함하는 2,277 nucleotide의 염기를 분석하였으며 gp64의 early, late 프로모터 발현을 조절하는 인자들을 확인하였다. gp64프로모터를 이용한 transient 발현 벡터를 제작하고 외래유전자로써 lacZ 유전자를 Bm5 세포주에서 transient 발현시켰다. 세포주 내에 도입된 플라스미드 DNA의 안정성을 확인하였으며, gp64 프로모터의 외래유전자 발현성 여부를 조사하기 위하여 gp64 프로모터 하에 laxZ 유전자를 가지는 재조합 바이러스를 제작하고 $\beta$-galactosidase in 냐셔 staining을 수행한 결과 전체적인 발현량은 매우 약한 것으로 판단되었다. BmNPV-K1의 gp64 프로모터를 이용한 벡터는 더욱 민감한 표지 유전자를 발현시켜 재조합 바이러스의 분리에 이용하거나 숙주세포에 독성을 보이는 유전자 산물의 소량 발현에 더욱 유용할 것으로 판단된다.

A strain of Actinomycetes producing cathepis B inhibitor was isolated from soil and identified as Streptomyces misakiensis. The product of S. misakiensis inhibited effectively cathepsis B proteinases as well as trypsin and papain. The cathepsin B inhibitor were largely produced with incubation for 4 days. The S. misakiensis was the most growth with incubation for 5 days. The cathepsin B inhibitor was isolated from the extraction of both with ethanol, ethanol and chlorofrom, and following several column chromatography such as sephadex G-15, silica gel 60 and sephadex LH-20 chromatography. The moleculer weight of purfied inhibitor was 138 dalton.

AA의 2번 위치의 수산기에 부위특이적 활성을 갖는 Paenibacillus sp. JB-13 유래 CGTase의 AA-2G 생산최적조건을 검토하였다. AA-2G 생산에 효율적인 당공여체를 조사하기 위해 다양한 포도당 중합체를 당공여체로 사용하여 AA-2G 생산성을 검토한 결과 dextrin이 가장 높은 AA-2G 생산성을 나타내었으며, 대체적으로 중합도가 높은 당을 효율적으로 이용하였다. 최적 당공여체인 dextrin을 사용하여 AA-2G 생산최적조건을 검토한 결과 반응 혼합액을 2,500 units/ml의 CGTase, 기질농도, 15%, 기질농도비(AA-g/dextrin-g) 3:2으로 조성하여, $37^{\circ}C$ , pH 6.5에서 44시간 반응 시킨 후, 1,500 units/ml의 glucoamylase를 $55^{\circ}C$에서 15시간 반응시켰을 때 AA-2G의 생산이 최대를 나타내었다.

우리 지역 토양환경에 도착해 살고 있는 토착 길항미생물을 분리, 선발, 육종하여 다시 지역모양으로 돌렸을 때 우점 능력이 크고 적응력이 큰 토착 생물방제균을 선발하고자 경북 경주지역에서 자연농법을 수행하고 있는 저병해경작지로부터 토착 길항미생물을 분리하였다. 이들 중 고추역병균 Phytophthora capsici의 생육을 강력히 길항하는 항생물질과 siderophore를 동시에 생산하는 복수 길항기작의 토착 길항세균 Pseudomonas fluorescens 2112를 선발하였고, 이 균주로부터 우선 항진균성 항생물질을 생산, 정제한 후 그 특성을 조사하였다. 이 항생물질은 산성 pH 조건에서는 매우 안정하였으나 알칼리성 용액에서는 불안정한 물질로 판단되었고, 또한 열에 비교적 안정하여 $80^{\circ}C$까지 30분동안 열처리하여도 항균활성을 50% 정도 유지하였다. P. fluorescens 2112로부터 이 균주의 중요 생물방제 기작인 항진균 항생물질을 단일물질 수준으로 분리하여 그 구조를 분석해 본 결과 선발된 균주가 생산하는 항생물질은 다른 Pseudomonas sp. 에서도 생산되는 것으로 알려진 2,4-diacetylphloroglucinol로 동정이 되었다. 고추역병균 P. capsici의 생육을 억제하는 토착 길항세균 P.fluorescens 2112가 생산하는 항진균 항생물질의 항균범위를 조사해 본 결과 다양한 식물병원성 진균에 방제력을 가지는 광범위 항생물질임을 확인 할 수 있었지만, Fusarium 속에 대해서는 큰 활성을 볼 수 있었다. 이 항진균 항생물질을 이용해 저해기작을 고추역병균 P. capsici의 포자를 대상으로 radioisotope labelling precursor [$^{3}$ H] Leucine, [$^{14}$ H] Glucose, [$^{3}$ H] Adenine 등으로 조사한 결과, 그 길항기작이 RNA 합성 저해로 추정되었다.

Indole and its derivatives form a class of toxic recalcitrant environmental pollutants, Abacte-rium, strain KL-9 was isolated from soil with indole as a sole source of carbon and nitrogen. KL-9 was identified as Acinetobacter sp. on the basis of 16 S rRNA gene sequence, fatty acid and quinone compositions. This identification was also confirmed by the ability of carbon source utilization and other biochemical tests. The growth of Acinetobacter sp. KL-9 was fastest with 0.3mg/ml of indole as was inhibited by higher than 0.5mg/ml of indole in the medium, KL-9 with indole also produced indigo. The formation of indigo was stimulated inthe presence of glucose, which is not a growth-suppoting carbon source for KL-9. Additional biotransformation evidence showed that anthranilate is an intermediate for the degradation of indole KL-9.

B. subtilis cx1이 생산하는 박테리오신(BSCX1)을 부분 정제하고 특성을 규명하였다. BSCX1은 pH 안정성 실험에서 pH 2.5~9.5 구간에서도 안정되게 항균활성을 유지하였다. 각종 효소에 대한 안정성 실험에서는 protease, trypsin, proteinase K, 그리고 carboxypeptidase로 처리하였을 때는 완전히 항균활성이 사라졌고, $\alpha$-amylase, lipase, aminopeptidase는 항균활성에 영향을 미치지 못하였다. 이상의 결과로 본 항균물질이 단백질계열의 물질이며 당이나 지질 결합은 항균활성에 큰 영향을 미치지 않는 것으로 추정되었다. 그러나 열에는 불안정하여 $60^{\circ}C$이상에서는 15분 안에 완전히 그 항균활성을 상실하였으며 $50^{\circ}C$ 에서는 15분만에 역가의 50%가 실활 되었다. 이에 반하여 저온에서는 무척 안정하여 $-20^{\circ}C$와 $-70^{\circ}C$에서 수개월간 보관하여도 여전히 항균활성을 그대로 유지하였다. Tricin-SDS-PAGE를 통하여 BSCX1의 분자량은 약 9,500 dalton으로 확인되었ekl. BSCX1은 pH2.5에서 9.5에 이르는 넓은 pH 영역에서 그 활성을 유지하므로, 기존에 개발된 nisin이나 유산균 유래의 항균물질이 pH 안정성이 떨어진다는 단점을 보완할 수 있는 생물학적 식품보존제로서의 개발 가능성을 높이 시사하였다.

For the development of probiotics in aquaculture of marine organisms, three strains having psychrophilic and salt tolerant characteristics were isolated from Kimchi. Amng the isolated strains. MG19, MG89 and MG208 were identified as Lactobacillus brevis, Enterococcus faecium and Lactobacillus plantarum, respectively. The neutralized culture broth of isolated strains were tested in order to evaluate the antibactrial activity, 조초 showed high antibacterial activity against Edwardsiella tarda, Vibrio cholerae andPseudomonas fluorescens. In mixed culture of pathogens and isolated strain,pathogens were signifi-cantly inhibited after 2 days of cultivation but the isolated strains showed normal growth. When the Edwardsiella tarda was cultured with three isolated strains, its growth was completely inhibited after 254 hours of cultivation. The effect of isolated three strains as probiotics was investigated based on the changes in body weight of aquacultured flounder. After 50 days feeding trial, it was found that the mean body weight gain of the tested group fed freeze-dried isolated cells was significantly greater than that of the control group. These results suggest that these isolated strains can play an important role as probiotics in aquaculture.

This study was performed to investigate antigenic expression patterns in the course of HSV-1 infection. In SDS-PAGE analysis, HSV-1 antigens were detected, and among them, antigens in the size of 39, 47, 63, 86, 101, 105, 135, 159, and 181 kDa appear to be expressed in the most dominant forms. BALB/c mice were infected with HSV-1 for 29 days and antigenic expression from HSV-1 was investigated by Western blot analysis using anti-HSV-1 sera collected every two days from BALB/c mice infected with HSV-1. Most of HSV-1 antigens appeared sporadically as the infection progressed. However, antigens in the sizes of 63kDa and 135kDa were expressed from day 1 and 3, respectively, and existed continuously during the course of infection for 29 days, suggesting that they are the most dominant antigens inducing immune response durign HSV-1 infection, and they could be the target antigens for the development of vaccines. The isotype levels of IgA, IgGl, and IgM increased till the 17 th day infection and then started to decrease. During this course. IgGl was the most dominant isotype. In an indirect immunofluorescent assay, antibodies exhibited surface binding to the Vero cell infected with HSV-1, demonstrating that HSV-1 antigens are expressed on the surface of Vero cells.