The studies were carried out to obtain the basic data for maximizing the protoplast yields from the mycelia of P. ostreatus and P. sajor-caju. Some factors affecting the regeneration of the protoplast of both species and the productivity of their reversion were also examined. The maximum yields of protoplasts were obtained from four days cultured mycelia of both species on cellophan membrane placed on the surface of PSA or MCM media in a petri dish. The optimal concentration of lytic enzyme Novozym 234 for protoplast releasing was 5 mg per ml of 0.5 M phosphate buffer solution with 0.6 M sucrose or 0.6 M $MgSO_4$ at pH 6.0. The greatest number of protoplasts was released 3 hours after incubation of the mycelia of P. ostreatus and after 4 hours for the P. sajor-caju in the lytic enzyme solution. Among the osmotic stabilizer solutions tested 0.6 M sucrose and 0.6 M KCl showed the best regeneration rates of the protoplasts of both species. When 0.75 % agar solution was over-layed on the regeneration media immediately after inoculation of the protoplast the regeneration rates were greatly enhanced. The ampicillin added to the agar solution prevented bacteria from infection. The reverted isolates produced the sporophores and basidial spores just like their parents without any mutations when they were cultivated in a broad mouth bottle with sawdust substrates.
This study was conducted to determine types of seed dormancy in Sambucus racemosa subsp. pendula (Nakai) H.I. Lim & C.S. Chang, an endemic tree species of Korea, whose seeds have been considered difficult to germinate. Seeds of S. racemosa subsp. pendula were stratified at 25/15 or $5^{\circ}C$ for 0, 6, or 12 weeks (wks) and incubated at 15/6, 20/10, 25/15, or $30/20^{\circ}C$ (12/12 h) under 14 h photoperiod. To determine the effect of $GA_3$ on seed germination of S. racemosa subsp. pendula, seeds were treated with 0, 500, or 1000 ppm $GA_3$ and then germinated at 25/15 or $5^{\circ}C$. The change in embryo length was investigated at 25/15 or $5^{\circ}C$. Seeds given 12 wks of cold stratification germinated to 33.4% at $15/6^{\circ}C$ and to 25.4% for seeds given 6 wks of warm stratification + 12 wks of cold stratification at $20/10^{\circ}C$. At $25/15^{\circ}C$, seeds given 12 wks of warm stratification + 6 wks of cold stratification germinated to 26.0%, and to 28.2% for seeds given 12 wks of warm stratification + 12 wks of cold stratification at $30/20^{\circ}C$. Warm stratification alone did not germinate seeds throughout the experiment, regardless of the thermoperiod. Linear embryos began to grow after 60 days of incubation at 25/15 or $5^{\circ}C$. The embryo length at day 69 increased from 1.4 mm to 1.50 or 1.62 mm at 25/15 or $5^{\circ}C$, respectively. Embryos of S. racemosa subsp. pendula seeds grew better at $5^{\circ}C$ than at $25/15^{\circ}C$. Gibberellin was effective to break seed dormancy of S. racemosa subsp. pendula. Seeds treated with 500 ppm $GA_3$ germinated up to 40.0% at $25/15^{\circ}C$ and to 62.7% for those treated with 100 ppm $GA_3$ at $5^{\circ}C$. With these results, seeds of S. racemosa subsp. pendula have both nondeep complex and intermediate complex morphophysiological dormancy.
To detect proper lytic assay conditions of the crude zumolyase from Arthrobacter luteus, effets of the various factors involved in the lytic system of Sacchromyces sake cultured with shaking in the malt extracts medium were investigated. The results are summarized as follows : 1. The susceptibilities of viable cells of S. sake from logarithmic growth phase to the lytic enzmye were much greater than those of the cells in lag and stationary phases. The cells cultured for 18 hr were the most susceptible to the enzyme. 2. Lytic activity of the enzyme toward the viable cells of S. sake was very low. It was, however, enhanced 4 folds of more by the pretreatment of the cells with 0.05 M sodium sulfite. 3. Lytic activity of the enzyme toward commercial baker's yeast cells was negligible, and the effect of sodium sulfite on the lysis of the cells also was nothing but a little. 4. The lyophilized cells of the baker's yeast showed more susceptibility to the lytic enzyme than viable cells of the yeast. No definite effect of sodium sulfite on the lysis of the lyophilized cells, however, was observed either baker's yeast of S. sake. 5. It appeared that the relationship between the reaction rate and the enzyme concentration on the lysis of the yeast cell walls followed enzyme kinetic theory, but one between the reaction rate and concentration of the yeast cells as substrates showed different pattern from that in enzyme kinetic theory. 6. After the preparation of crude zymolyase was kept at $7^[\circ}C$ for 10 days in the 0.05 M phosphate buffer, pH 7.5, the remainning lytic activity was about 80 %.
The objective of this study was to examine if transient transfection of CAR can transactivate CYP2B1 PBRU reporter gene in COS cells in which the endogenous CYP2B1 gene is not induced by PB. In non-transfeced cells of both Hep G2 and COS, the endogeneous expression of CAR was not detected by antibody against CAR. When cultured cells were transfected with CAR expression plasmid, mCAR1-GFP, both cell types expressed high levels of CAR protein and could allow to examine the effect of CAR in PBRU transactivation. Both cell types expressed endogenous RXR and transfection of RXR expression plasmid dramatically increased its protein expression. Whereas CAR transactivated PBRU2C1Luciferase about 12 fold as compared to 2C1Luciferase in Hep G2 cells, it did not stimulate the luciferase activity of the PBRU reporter gene in COS cells. These results indicate that Hep G2 cells can respond to CAR differently from COS cells, and suggest that factors other than CAR and RXR may be required in inducing PBRU activation and the expression of these factors may be different between liver and kidney.
Hyun Bek;Lim Hak-Seob;Chung Kyung Kae;Choi Yung Hyun;Choi Byung Tae;Seo Min-Jeong;Kim Ji-Eun;Ryu Eun-Ju;Huh Man Kyu;Joo Woo Hong;Jeong Young Kee
Journal of Life Science
/
v.15
no.6
s.73
/
pp.987-993
/
2005
A bacterium, producing a fibrinolytic enzyme, was screened from a decaying rice plant. The bacterium was identified as Bacillus subtilis by morphological, biochemical, and physiological properties and named Bacillus subtilis BK-17. The fibrinolytic enzyme (BK) was purified from supernatant of Bacillus subtilis BK-17 culture broth. The molecular weight was 31 kDa as determined by SDS-PAGE. The effect of temperature, pH, and plasminogen on the activity of the bacillokinase (BK) was analysed and the activity was compared with urokinase. The optimal temperature and pH were $50^{circ}C$ and pH 7, pH 8, respectively. The BK activity was inhibited to $45\%$, $35\%$, and $23\%$ with 1mM EDTA, $Zn^{2+}$, and $Ca^{2+}$, respectively. However, $Mg^{2+}$, $Mn^{2+}$, and $Co^{2+}$ ions did not have any significant effect on the enzyme activity The BK showed the artivity in the both plates, plasminogen-free fibrin plate and plasminogen-rich fibrin plate. The result indicates that the BK can directly act the fibrin. In comparison of fibrinolytic activity with urokinase on the fibrin plate, the BK shows about 20 folds higher activity than that of the urokinase.
Ryu, Myeong Seon;Yang, Hee-Jong;Jeong, Su-Ji;Seo, Ji Won;Ha, Gwangsu;Jeong, Seong-Yeop;Jeong, Do-Youn
Korean Journal of Microbiology
/
v.54
no.4
/
pp.384-397
/
2018
The aim of this study is to screen the strains of Bacillus spp. possessing safety, probiotic activity, and so on, which can be utilized as probiotic resource for using the feed and supplement food of companion animal. About 300 isolates were isolated from traditional Korean sauces, four isolates that did not have or produce the six kinds of B. cereus type vomiting and diarrhea toxin genes, ${\beta}$-hemolytic, and three kinds of carcinogenic enzymes were selected. Antibiotic gene retention, cell surface hydrophobicity, antibiotic sensitivity, and glucose utilization were analyzed for four isolates, and finally SRCM 100731 was selected. SRCM 100731 was named as Bacillus amyloliquefaciens SRCM 100731 16S rRNA sequencing analysis, and carried out optimization of cell growth for industrial applications such as pet food and feed. The effects of 14 different components on cell growth were investigated and three significant positive factors, molasses, sodium chloride, and potassium chloride were selected as the main factors based on a Plackett-Burman design. In order to find out optimal concentration on each constituent, we carried out central composite design. The predicted optimized concentrations were 7% molasses, 1.1% sodium chloride, 0.5% potassium chloride. Finally, an overall about 7-fold increase in dry cell weight yield ($12.6625{\pm}0.0658g/L$) was achieved using the optimized medium compared with the non-optimized medium ($1.8273{\pm}0.0214g/L$). This research is expected to be highly utilized in the growing pet industry by establishing optimal cultivation conditions for industrial application as well as screening Bacillus amyloliquefaciens SRCM 100731 as probiotic resource for companion animal.
A microbial insecticide "Bt-Plus" has been developed to enhance an insecticidal efficacy of an entomopathogenic bacterium, Bacillus thuringiensis (Bt). However, its wettable powder formulation is not preferred by farmers and industry producers due to relatively high cost. This study aimed to develop a soluble concentrate formulation of Bt-Plus. To this end, an optimal mixture ratio of two bacterial culture broths was determined to be 5:4 (v/v) of Bt and Xenorhabdus nematophila (Xn) along with 10% ethanol preservative. In addition, Bt broth was concentrated by 10 times to apply the mixture at 1,000 times fold dilution. The resulting liquid formulation was sprayed on cabbage crop field infested by late instar larvae of the diamondback moth, Plutella xylostella. The field assay showed about 77% control efficacy at 7 days after treatment, which was comparable to those of current commercial biopesticides targeting P. xylostella. For storage test in both low and room temperatures, the liquid formation showed a relatively stable control efficacy at least for a month. To develop a quality control technique to exhibit a stable control efficacy of Bt-Plus, Bt spore density ($5{\times}10^{11}$ spores/mL) and eight active component concentrations of Xn bacterial metabolites in the formulation products have been proposed in this study.
Bak, Won Chull;Lee, Eun Young;Yoon, Kab He;Lee, Won Kyu;Yi, Chang Keun;Hong, Ki Sung
Journal of Korean Society of Forest Science
/
v.83
no.1
/
pp.12-19
/
1994
Studies were made to find out the mycelial growth and rotting characteristics of 91 Lentinus edodes isolates collected or hybridized in Korea and abroad, and their adaptability to bag-culture. The results are as follows. 1. There were little differences in mycelial growth at PDA and oak (Quercus mongolica) sawdust-based medium, and rotting among high-, mid- and low temperature types of L. edodes isolates. But, the fruit yield of high-temperature type was much higher than those of the other two types. The fruit yields of the isolates in the same strain group were apparently different. 2. From the sawdust-based culture of 91 isolates, FRI 221, FRI 208 and FRI 169 were selected as excellent strains in yield and Quality, showing fruiting of 157g, 152g and 119g per 1kg-medium, respectively. 3. Attempts to culture in various media with different substrates resulted in almost same fruit yields, and the yield was proportional to medium weights as 2.5kg-medium showed 2.5 times more yield than 1kg-medium. Fruit yields were different according to incubation periods and the period of little more than 100days was best. 4. When the correlation analyses among mycelial growth, rotting ability, yield and fruit-shape normality were made with the 91 isolates, mycelial growth at sawdust-based medium showed highly positive correlation with that at PDA, while fruit yield was negatively correlated with mycelial growth in PDA and sawdust-based medium.
To study the behavior of $NH_4{^+}$ of CMS (condensed molasses solubles) in soil, a laboratory incubation experiment was conducted during a period of up to 21 days at $25^{\circ}C$. The $NH_4{^+}$ of CMS was labeled with $^{15}N$ and was applied to water-unsturated and water-saturated conditions. Soil pH was gradually decreased from 6.1 to 5.4 under unsaturated condition. However, soil pH was increased to 6.5 within 2 days under saturated condition and then was constant. The concentration of ammonium was decreased 3 times faster under unsaturated condition than under saturated condition. The concentration of nitrate was increased from 17.4 to $155.4mg\;kg^{-1}$ under unsaturated condition. But concentration of nitrate was kept with low(below $8.0mg\;kg^{-1}$) under saturated condition. During the incubation, 52.4% of $^{15}NH_4{^+}$ applied was existed in the form of $NO_3{^-}$ by nitrification under unsaturated condition. Most of applied nitrogen was immobilized within 4 days of incubation. On 21 days of the incubation the percentage of immobilized nitrogen derived from $^15NH_4{^+}$(NDFA) was 19.6% under unsaturated condition and 17.0% under saturated condition. The percentage of unaccounted N, which was lost by denitrification, was 28.4% under unsturated condition and 67.6% under saturated condition.
The low seed propagation is one of the problem needed a lot of seed tuber for the propagation in yam. Therefore this experiment was carried out to understand the possiblility of seed tuber propagation by tissue culture of yam. In-vitro stem node of yam was cultured by concentration treatments of 1/2, 1/4 and 1/8 with MS medium additted with each concentration levels of IAA, NAA, IBA, kinetin and BA. Acorrding to the Iower concentration than MS medium, length of shoots was promoted, leaf emergence shoots and rooting shoots were increased at 1/8 MS medium during the culturing period of stem node in yam. Fixed IBA and kinetin under the concentration of MS mdeium was inhibited severely by the heigh concentration additted with lAA $1mg\;/\;{\ell}\;and\;NAA\;4mg\;/\;{\ell}$. But fixed IBA $5mg\;/\;{\el}l\;and\;kinetin\;2mg\;/\;{\ell}$ with concentration of 1/8MS medium was remarkably promoted leaf emergence shoots and rooting shoots by $1mg\;/\;{\ell}$ of additted lAA and NAA. Percentage of induced shoots was increased by combination treatments of lAA. $1.5mg\;/\;{\ell}\;and\;kinetin\;2mg\;/\;{\ell}$, also leaf emergence shoots and rooting shoots were promoted by combination treatments of lAA $1.5mg\;/\;{\ell}\;and\;kinetin\;2mg\;/\;{\ell}$.
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