Two putative regulator genes, mocR and mocS, of the moc (mannityl opine catabolism) operons in pTi15955 of the octopine-/mannityl opine-type Agrobacterium tumefaciens strain 15955, were tested for their possible roles as repressors in the moc operons. The regions upstream of macC and mocD, the first structural genes in the two divergently oriented moc operons, were transcriptionally fused into the promoterless lacZ reporter gene. Each of the lacZ-fusions was introduced into Agrobacterium strain UIA5, a Ti plasmid-cured derivative, harboring either a mocR or a mocS clone. The resulting strains were grown in media containing various sugar sources, and the $\beta$-galactosidase activities were quantitatively measured. The results suggested that MocR repressed the expression of macC and macD. The expression of the fused $\beta$-galactosidase was not induced by mannopine (MOP) or possible catabolic intermediates of the opine, e.g. santhopine (SOP), glucose, mannose, or glutamine. However, the repression was significantly relieved by the supplementation of MOP and the concomitant introduction of the agcA gene encoding MOP cyclase that catalyzes the lactonization of MOP to agropine (AGR). These results suggested that AGR, rather than MOP or the other catabolic intermediates, is the inducer for the expression of the operon. On the contrary to previous report showing that the induction levels of macC and macD were lowered by the supplementation of inorganic nitrogen in media, the expression of these genes was not affected by the level of nitrogen in our reporter system. MocS did not strongly repress the expressions of macC and mocD. It is possible that MocS may be involved in the regulation of the operons present downstream of the moc operon, which are responsible for the utilization of mannopinic acid and agropinic acid.
Lee, Bae Ik;Kim, Shin Kwon;Kim, Jong Hyeok;Kim, Hyung Seop;Kim, Jong Im;Shin, Woongghi;Rho, Jung-Rae;Yih, Wonho
ALGAE
/
v.34
no.2
/
pp.163-175
/
2019
To compare the nutritional quality of TPG (Teleaulax / Plagioselmis / Geminigera) clade species of cryptomonads with that of RHO (Rhodomonas / Rhinomonas / Storeatula) clade species 6 Teleaulax amphioxeia (TA) and 1 Rhinomonas sp. strains were mass-cultured in newly designed 500-L photobioreactors to the end of exponential growth phase. Intraspecific variations (IVs) in terms of one standard deviation among the 6 TA strains in the compositions of the three macronutrients were 41.5 (protein), 89.8 (lipid), and 15.6% (carbohydrate) of the mean. When harvested from stationary growth phase mean compositions of essential amino acids (EAAs, 47.3%) and non-EAAs (52.7%) of the 2 TA strains, CR-MAL07 and CR-MAL08-2, were similar to those of a Chroomonas strain. The IVs between the 2 TA strains in the composition of EAAs (10.3 and 2.4) and non-EAAs (8.5 and 2.1% of the mean) were rather smaller than those of saturated fatty acids (30.3 and 26.1) and unsaturated fatty acids (UFAs, 12.0 and 12.5% of the mean) in f/2-Si and urea-based compound fertilizer (UCF) culture media, respectively. Mean compositions of eicosapentaenoic acid (EPA, 17.9%) and docosahexaenoic acid (DHA, 12.7%) of total fatty acids of the 2 TA strains were higher than those that of a Chroomonas strain. EPA and DHA compositions exhibited similar level of IVs between the 2 TA strains in f/2-Si (14.6 and 11.0) and UCF media (12.6 and 13.5% of the mean). Thus, the nutritional quality in terms of amino acids, UFAs, EPA, and DHA in a TPG clade species, T. amphioxeia was comparable to those of RHO clade species with notable IVs. Practically, biotechnological targets for TPG clade cryptomonad strains might be subspecies or clone level.
Chung, Kun-Sub;Rafael, F.R.;Oh, Sang-Suk;Richardson, T.
Journal of Microbiology and Biotechnology
/
v.1
no.1
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pp.31-36
/
1991
Yeast expression plasmids containing an appropriate leader sequence and bovine $\beta$-casein cDNA were constructed to produce $\beta$-casein for the study of its functional characteristics. Two kinds of expression systems for $\beta$-casein were constructed using pCGYl444 as a precursor plasmid. This plasmid is a yeast-E. coli shuttle vector which contains the chelatin promoter. The plasmid pISB202 contains the invertase leader sequence and $\beta$-casein gene. The plasmid pDEB303 contains the original bovine $\beta$-casein leader sequence gene. These two plasmids were introduced into S. cerevisiae AB116 which is a strain deficient in the major yeast proteinases. Each clone was grown in minimal media for 24 h before induction by $CuSO_4$. The cells were thus grown under expression conditions. Both strains harbouring pISB202 and pDEB303 expressed bovine $\beta$-casein. The $\beta$-casein was detected using immunochemical staining after western blot. Secretion of $\beta$-casein was detected in the culture broth. The estimated amount of secreted $\beta$-casein was approximately 50 ${\MU}g$/l.
SMMIAR (Submerged Moving Media Intermittent Aeration Reactor) Process is a very efficient system which remove ammonia to nitrogen gas in one reactor. In this study, we determined the diversity of ammonia oxidizing bacteria and denitrifying bacteria using specific PCR amplification and the clone library construction. An ammonia monooxygenase gene(amoA) was analyzed to investigate the diversity of nitrifiers. Most of amoA gene fragments (27/29, 93%) were same types and they are very similar (>99%) to the sequences of Nitrosomonas europaea and other clones isolated from anoxic ammonia oxidizing reactors. ANAMMOX related bacteria have not determined by specific PCR amplification. A nitrite reductase gene(nirK) was analyzed to investigate the diversity of denitrifying bacteria. About half (9/20, 45%) of denitrifiers were clustered with Rhodobacter and most of others were clustered with Mesorhizobium (6/20, 30%) and Rhizobium (3/20, 15%). All of these nirK gene clones were clustered in alpha-Proteobacteria and this result is coincide with other system which also operate nitrification and denitrification in one reactor. The molecular monitoring of the population of nitrifiers and denitrifiers would be helpful for the system stabilization and scale-up.
The monoclonal antibody to chloramphenicol(CAP) was produced to develop an enzyme-linked immunosorbent assay(ELISA) for residual CAP. An immunogen(CAP-BSA) was prepared by immunogen, antibody titer was measured by indirect ELISA. Spleen cells form the immunized mouse were fused with SP2/OAg14 myeloma cells. Among hybridomas selected in HAT media, 6 clones shown high antibody titer to CAP were subjected to cloning by limit dilution, and all of the monoclonal antibodies(MCA1, 2, 3, 4, 5, 7 and 9) produced by each clone were identified as IgG1 by ELISA isotyping analysis. Competitive ELISA condition was established by using the purified monoclonal antibody MCA1 as primary antibody and CAP-HSA conjugate as coating antigen. Standard curve of CAP(n=28) showed that the lowest detection limit of CAP is 20ng/ml level. The cross-reactivities of the 6 monoclonal antibodies showed that CAP sodium succinate. CAP base, P-nitrophenol, and p-nitrobenzyl alcohol were 89∼178, 0.050∼2.237, 0.056∼0.794 and 0.013∼7.939%, respectively. No cross-reactivities were observed with phenylalanine, tyrosine, glutamine, thiamphenicol, neomycin, streptomycin, gentamicin, sulfamethazine, sulfathiazole, chlortetracycline and p-aminobenzoic acid.
Present study describes a method on the application of efficient tissue culture systems for the micro-propagation of juvenile and mature sawtooth oak(Quercus acutissima). Nodal segments with axillary buds were used as initial explant sources. WPM(Woody Plant Medium) was the best in growth and proliferation of shoot among the media tested. Although the single effect of zeatin revealed on two dorminant shoot elongation with normal growth until the elevation of levels up to 3.0mg/l, BAP($N^6$-benzyl amino purine) usually showed better response than zeatin on shoot multiplication and/or elongation. In addition, the incorporation of BAP and zeatin onto the culture media represents more effectiveness in shoot proliferation and its growth. Optimum concentrations of BAP and zeatin were 0.5 and 0.05~1.0mg/l, respectively. Ninety percent of the proliferated shoots was rooted on half-strength GD (Gresshoff and Doy, 1972) medium containing 0.5mg/l IBA(indole butyric acid) in 4 weeks after culture. More than 70% of the rooted plantlets survived after 5 months of transplanting into artificial soil mix containing equal amount of peatmoss and perlite. Among 27 plus tree clones which were grafted twice onto the juvenile rootstocks, only 4 clones revealed the possibility for shoot multiplication through tissue culture system. The capacity for the micropropagation using mature explant sources was highly depended on clonal differences compared with those of octet age. More than 90% of rooting ratio was obtained from the best responding clone. Among the 7 rooting media tested, GD medium was the best far rooting. The most effective rooting was obtained on half-strength GD medium containing 0.2 to 2.0mg/l IBA. More than 60% of rooted plantlets survived after 5 months of transplanting into the artificial soil mix.
Printmaking is a field of fine arts and is placed on a vague boundary that is perceived as a commercial product with a popular character due to the speciality of editions. Advances of modern science and technology has developed a new technique of printmaking, and the fusion of printmaking and computer has shown the possibility of reproduction art. Reproduction printmaking has been heavily influenced by photography and extended areas, and the various forms of printing have brought about many changes and attempts by stimulating the possibilities of indirect art at various angles. As the history of printmaking and technology closely relate, the development of computer makes widespread expansion of plural artistry, technological and artistic change. A new conceptual shape can be created on the copied image simply by placing the material of the print on the smoothly flat surface expressed in digital form. The process and the result of such work show the area of unique work which is different from the value of the $\grave{a}$ la carte art or the characteristics of the material given by the print. The deprecated perception of reproducibility evaluated the value of the work in a direct sense of printmaking. It is undeniable that it is devalued by a bundle of works regardless of the value of each edition. However, the physical properties of the prints on the paper are brought up with hand drawings drawn on the canvas by hand. And it becomes an opportunity to show new aspect and change through the process of combining digital print information on paper. The diversity of media is sometimes a controversy of identity between art and technology. In the future, it should be discussed how the limit of the media which can be enjoyed in the field of art can be set as a standard.
K. C. Hwang;D. W. Ok;D. N. Kwon;H. K. Shin;Kim, J. H.
Proceedings of the KSAR Conference
/
2001.03a
/
pp.52-52
/
2001
Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic Many nucleoside diphosphate (NDP) kinases are ubiquitous enzymes responsible for the exchange of ${\gamma}$-phosphates between tri- and diphosphonucleosides. The catalytic reaction follows a ping-pong mechanism in which the enzyme is transiently phosphorylated on a histidine residue conserved in all nucleoside diphosphate kinases. Beside their role in nucleotide synthesis, these enzymes present additional functions, possibly independent of catalysis, in processes such as differentiation, cell growth, tumor progression, metastasis and development. To clone murine nm23-M5, several expressed sequence tags (ESTs) of the GenBank data base, selected according to their homology to nm23-H5 cDNA, reconstituted a complete open reading frame (GenBank AF222750). To test whether murine NDPKs (1, 2, 3, 4, 5, and 6) can inhibit Bax-mediated toxicity in yeast, co-transformation was performed respectively. The yeast S.cerevisiae was transformed with a copy expression plasmid containing the histidine selection marker and expressing murine Bax under the control of a galactose-inducible promoter. Several clones were selected and found to be growth inhibited when Bax expression was induced with galactose. A representative clone was transformed again with a copy expression plasmid containing the tryptophane selection marker and expressing either murine Bcl-xL or NDPK under the control of a galactose-inducible promoter. Several subclones of the double-transformants were selected and characterized. The ability of Bcl-xL and NDPKs to suppress Bax-mediated toxicity was determined by growing yeast cells overnight in galactose media and spot-testing on galactose plates starting with an equal number of yeast cells as determined by taking the OD$_{600}$. Ten-fold serial dilutions were used in the spot-test. Plates were grown at 3$0^{\circ}C$ for 2-3 days. All murine NDPKs suppressed Bax dependent apoptosis. Futher study will be peformed whether Bax-toxicity inhibition was caused by NDP kinase activity or additional function.n.
Won Mi-Kyoung;Park Chan-Jin;Chang Kyoung-Soo;Kim Chang-Whe;Kim Yung-Soo;Isa Zakiahbt Mohd;Ariffin Yusnidar Tajul
The Journal of Korean Academy of Prosthodontics
/
v.41
no.6
/
pp.720-731
/
2003
Statement of problem. The importance of fixture design and surface treatment. Purpose. The clinical success of dental in plants is affected by many factors such like as degree of osseointegration, the effective load dispersion for the prostheses, and a lot of attempts have been made to overcome the difficulties. In this study, efforts were made to find the possibility of clinical acceptance of the dental implants of newly designed surface and resorbable blast media surcace. Materials and methods. In this study, two groups of custom-made, screw-shaped implants were prepared. The first with the consisting of Branemark clone design and the other with the new design. These implants were divided into four groups according to the kinds of surface treatment. Four implants($AVANA^{(R)}$, Osstem, Busan, Korea)of each group were installed in twenty rabbits. Group A was consisted of Branemark done implant left as machined, Group B with Branemark clone implants with RBM(Resorbable blast media) surface, Group C with newly designed implants left as machined and Group D with newly designed implants with RBM surface. One of the twenty rabbits died from inflammation and the observation was made for six weeks. Specimens from four groups were observed using scanning electron microscopy with 40, 100, 1000 magnification power and microsurface structures were measured by white-light scanning interferometry for three dimensional surface roughness measurements(Accura $2000^{(R)}$, Intek-Plus, Korea.). Removal torque was measured in 17 rabbits using digital torque gauge(MGT 12R, Mark-10 corp., NY, U.S.A.) immediately after the sacrifice and two rabbits were used for the histologic preparation(EXAKT $310^{(R)}$, Heraeus Kulzer, wehrheim, Germany) of specimens and observed under light microscope. Resonance frequency measurement($Osstell^{(R)}$) was taken with the 19 rabbits at the beginning of the implant fixation and immediately after the sacrifice. Results. Following results were taken from the experiment. 1. The surface of the RBM implants as seen with SEM had rough and irregular pattern with reticular formation compared to that of fumed specimens showing different surface topographies. 2. The newly designed implant with RBM surface had high removal torque value among four groups with no statistical significance. The average removal torque was $49.95{\pm}6.70Ncm$ in Group A, $51.15{\pm}4.40Ncm$ in Group B, $50.78{\pm}9.37Ncm$ in Group C, $51.09{\pm}4.69Ncm$ in Group D. 3. The RFA values were $70.8{\pm}4.3Hz$ in Group A, $71.8{\pm}3.1Hz$ in Group B, $70.9{\pm}2.5Hz$, $72.7{\pm}2.5Hz$ in Group D. Higher values were noted in the groups which had surface treatment compared to the untreated groups with no statistical significance. 4. The results from the histomorphometric evaluation showed a mean percentage of bone-to-implant contact of $45{\pm}0.5%$ in Group A, $55{\pm}3%$ in Group B, $49.5{\pm}0.5%$ in Group C, and $55{\pm}3%$ in Group D. Quite amount of newly formed bone were observed at the surface RBM-treated implants in bone marrow space.
To maintain its functional integrity, bone is continuously remodelled by a process involving resorption by osteoeclasts and formation by osteoblasts, In order to respond to changes in the physical environment or to trauma with the relevant action, this process is strictly regulated by locally synthesized or systemic fators, Prostaglandin $E_2(PGE_2$) is perhaps one of the best studied factors, having been known to affect bone cell function for several decades.$PGE_2$ has both anabolic and catabolic activities. Excess of $PGE_2$ has been implicated in a number of pathological states associated with bone loss in a number of chronic inflammatory conditions such as periodontal disease and rheumatoid arthritis. $PGE_2$ and other arachidonic acid metabolites have been shown to be potent stimulators of osteoclastic bone resorption in organ culture. The anabolic effects of $PGE_2$ were first noticed when an increase in periosteal woven bone formation was seen after the infusion of $PGE_2$ into infants in order to prevent closure of the ductus arteriosus. The cellular basis for the catabolic actions of $PGE_2$ has been well characterized. $PGE_2$increases osteoclast recruitment in bone marrow cell cultures. Also $PGE_2$ has a direct action on osteoclast serving to inhibit activity and can also indirectly activate osteoclast via other cells in the vicinity, presumably osteoblast. The cellular mechanisms for the anabolic actions of $PGE_2$ are not nearly so well understood. The purpose of this paper was to study the effects of $PGE_2$ and dibutyl(DB)cAMP on osteoblastic clone MC3T3El cells and on the generation of osteoclasts from their precursor cells. The effect of $PGE_2$ and DBcAMP on the induction of alkaline phoaphatase(AlP) was investigated in osteoblastic clone MC3T3El cells cultured in medium containing 0.4% fetal bovine serum. $PGE_2$ and DBcAMP stimulated ALP activity and MTT assay in the cells in a dose-dependent manner at concentrations of lO-SOOng/ml. Cycloheximide, protein synthesis inhibitor, inhibited the stimulative effect of $PGE_2$ and DBcAMP on ALP activity in the cells. $PGE_2$also increased the intracellular cAMP content in a dose-dependent fashion with a maximal effect at 500ng/ml. The effect of $PGE_2$ on the generation of osteoclasts was investigated in a coculture system of mouse bone marrow cells with primary osteoblastic cells cultured in media containing 10% fetal bovine serum.After cultures, staining for tartrate-resistant acid phosphatase(TRAP)-marker enzyme of osteoclast was performed. The TRAP(+) multinucleated cells(MNCs), which have 3 or more nuclei, were counted. More TRAP(+) MNCs were formed in coculture system than in control group. $PGE_2(10^{-5}10^{-6}M)$ stimulated the formation of osteoclast cells from mouse bone marrow cells in culture. $PGE_2(10^{-6}M)$ stimulated the formation of osteoclast cells from mouse bone marrow cells in coculture of osteoblastic clone MC3T3E1 cells This results suggest that $PGE_2$ stimulates the differentiation of osteoblasts and generation of osteoclast, and are involved in bone formation, as well as in bone resorption.
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