• Asian-Australasian Journal of Animal Sciences
• /
• v.12 no.1
• /
• pp.77-83
• /
• 1999

Molecular biological techniques that recently developed, have made it possible to realize some of new attempts in the research field of rumen microbiology. Those are 1) cloning of genes from rumen microorganisms mainly in E. coli, 2) transformation of rumen bacteria and 3) ecological analysis with nonculturing methods. Most of the cloned genes are for polysaccharidase enzymes such as endoglucanase, xylanase, amylase, chitinase and others, and the cloning rendered gene structural analyses by sequencing and also characterization of the translated products through easier purification. Electrotransformation of Butyrivibrio fibrisolvens and Prevotella ruminicola have been made toward the direction for obtaining more fibrolytic, acid-tolerant, depoisoning or essential amino acids-producing rumen bacterium. These primarily required stable and efficient gene transfer systems. Some vectors, constructed from native plasmids of rumen bacteria, are now available for successful gene introduction and expression in those rumen bacterial species. Probing and PCR-based methodologies have also been developed for detecting specific bacterial species and even strains. These are much due to accumulation of rRNA gene sequences of rumen microbes in databases. Although optimized analytical conditions are essential to reliable and reproducible estimation of the targeted microbes, the methods permit long term storage of frozen samples, providing us ease in analytical work as compared with a traditional method based on culturing. Moreover, the methods seem to be promissing for obtaining taxonomic and evolutionary information on all the rumen microbes, whether they are culturable or not.

• Journal of Nutrition and Health
• /
• v.29 no.1
• /
• pp.41-49
• /
• 1996

Bacterial transformation of bile acids is possibly involved in colorectal carcinogenesis. n several epidemiological studies, the fecal bile acid dietary fiber are related to the indicence of colonic cancer. This study investigated the influence of age, dietary fiber intake on fecal bile acid profiles in healthy subject. The dietary fiber were assessed by mean of 24 hour dietary recall method, the subjects consist of 238 members aged 20 to 64 years old and feces are collected from the subjects. Fecal bile acids and neutral sterols were analyzed by gas chromatography. Mean dily crude fiber intake level was 7.7$\pm$1.4g(dietary fiber : 16.7$\pm$3.5g), dietary fiber intake range being 6.5-36.8g. The dietary fiber intake in elederly subject was significantly lower than in the other younger groups. Dietary fiber intakes was negatively correlated with the total bil acid concentation in feces. Probably, a decrease in dietary fiber intake results in higher fecal bile acid concentrations. The secondary bile acid concentration is related to the colon cancer, deoxycholic acid and lithocholic acid were significantly higher in elderly subjects. Concentration of fecal total bile acid, deoxycholic acid, coprostanol, coprostanone were higher in low dietary fiber intake group. These results suggest that the risk factor for colon cancer might be reduced, when dietary fibers are consummed more.

• Economic and Environmental Geology
• /
• v.33 no.5
• /
• pp.353-365
• /
• 2000

After their first appearance on Earth, bacteria have exerted significant influence on geochemical behavior of elements. Numerous evidence of their control on geochemistry through geologic history has been observed in a variety of natural environments. They have mediated weathering rate, formation of secondary minerals, redox transformation of metals and metalloids, and thus global cycling of elements. Such ability of bacteria receives so considerable attention from microbiologists, mineralogists, geologists, soil scientists, limnologists, oceanographers, and atmospheric scientists as well as geochemists that a new and interdisciplinary field of research called 'geomicrobiology' is currently expanding. Some recent subjects of geomicrobiology which are studied extensively are as follows: 1) Functional groups distributed on bacterial cell walls adsorb dissolved cations onto cell surfaces by electrostatic surface complexation, which is followed by hydrous mineral formation. 2) Dissimilatory metal reducing bacteria conserve energy to support growth by oxidation of organic matter coupled to reduction of some oxidized metals and/or metalloids. They can be effectively used in remediating environments contaminated with U, As, Se, and Cr. 3) Bacteria increase the rate of mineral dissolution by excreting proton and ligands such as organic acids into aqueous system. 4) Thorough investigation on the effects of biofilm on geochemical processes is needed, because most bacteria are adsorbed on solid substrates and form biofilms in natural settings.

• 한국생물공학회:학술대회논문집
• /
• 2001.11a
• /
• pp.99-102
• /
• 2001

Recombinant Saccharomyces cerevisiae strains harboring various copy numbers of hirudin gene were developed to study dependency of hirudin expression level on its gene copy number. A linear relationship between the copy number of hirudin expression cassette and hirudin expression level was observed up to 10 copies. A double <5-integration vector truncated wi 디 1 the unnecessary bacterial genes before yeast transformation showed a four-fold increase in transformation efficiency and a 1.3-fold enhancement in hirudin expression level compared with a single <5 system. Gratuitous hirudin expression strain was developed by disrupting the GALl gene of S. cerevisiae. Glucose that was fed in a limited manner effectively supported cell growth and hi겨din expression by the gratuitous strain. Effects of methanol concentrations on hirudin production in recombinant Hansenula polymorpha were investigated in continuous and fed-batch cultures. At a steady-state of continuous culture, an optimum methanol concentration of 1.7 g/L was determined at a dilution rate of 0.18 $h^{-1}$ with 1.8 mg/L ${\cdot}$ h hirudin productivity.

• The Plant Pathology Journal
• /
• v.21 no.1
• /
• pp.39-46
• /
• 2005

The promoter region flanking the 5’ CAZFP1 coding region was isolated from the genomic DNA of Capsicum annuum. To identify the upstream region of the CAZFP1 gene required for promoter activity, a series of CAZFP1 promoter deletion derivatives was created. Each deletion construct was analyzed by Agrobacterium-mediated transient transformation in tobacco leaves after infection by Pseudomonas syringae pv. tabaci, or treatment with methyl jasmonate (MeJA), ethylene, abscisic acid (ABA), salicylic acid (SA), cold and wounding. Promoter fragments of 685 bp or longer showed 7-fold or greater induction after P. s. pv. tabaci infection and MeJA treatment. The CAZFP1 full-length promoter (-999 bp) also showed 6-fold induction in response to ethylene. The transiently transformed tobacco leaves with the CAZFP1 full length promoter fused-GUS gene showed more than 5-fold induction in response to SA, ABA and cold. These results suggest that the CAZFP1 promoter contains responsive elements for pathogen, MeJA, ethylene, SA, ABA and cold.

• Microbiology and Biotechnology Letters
• /
• v.40 no.3
• /
• pp.197-207
• /
• 2012

This study investigated the role of plasmids and their relationship with the multiple antibiotic resistance of 30 Vibrios sp. isolated from molluscs and crustaceans sampled from the Kerala coastal waters of India. The biochemical identification and antibiotic resistance profiles were determined, followed by the plasmid profiles, conjugation and transformation efficiencies. The results showed a considerable difference in the level of bacterial resistance to various antibiotics; while all 30 strains were found to be MAR Vibrios sp. and their resistance patterns varied. All the strains were resistant to amoxycillin, ampicillin and carbeniciliin. 87% were resistant to rifampicin; 74% to cefuroxime; 67 to streptomycin; 53% to norfloxacin and ciprofloxacin and 47% to furazolidone and nalidixic acid. In addition to their antibiotic resistance, the plasmid DNA of the MAR Vibrios strains isolated from the molluscs and crustaceans was also studied. Nine strains isolated from crustaceans and molluscs were found to harbor 1-3 plasmids with sizes varying from 5. 98 kb to 19. 36 kb. The average transformation efficiency was about $5{\times}10^{-8}$ and the conjugation efficiency varied from $2.1{\times}10^{-3}$ to $10^{-9}$. A further study of antibiotic resistance patterns may be useful to test the extent of drug resistance in seafoods and help to devise a nationwide antibiotic policy.

• KSBB Journal
• /
• v.4 no.2
• /
• pp.94-98
• /
• 1989

In vitro attachment experiments of bacteria to surface of host plant cell were carried out using C14 labeled cells of A. rhizogenes strain A4 and carrot protoplasts isolated from suspension culture of cells. Protoplasts were cocultivated with A. rhizogenes at various times after their isolation. Attachment kinetics showed that adherence of bacteria to protoplasts attained a maximum level within 120mins of co-cultivation. Maximum attachment occured at pH 6.0 and 24-35$^{\circ}C$. Bacterial attachment was observed at botg carrot cells with and without primary cell wall. The inhibition of transformation on the carrot root discs by A. rhizogenes was observed when non-related strain and heat inactivated bacterial strain cells were pretreated.

• Parasites, Hosts and Diseases
• /
• v.49 no.4
• /
• pp.349-356
• /
• 2011

The existence of symbiotic relationships between Acanthamoeba and a variety of bacteria is well-documented. However, the ability of Acanthamoeba interacting with host bacterial pathogens has gained particular attention. Here, to understand the interactions of Escherichia coli K1 and E. coli K5 strains with Acanthamoeba castellanii trophozoites and cysts, association assay, invasion assay, survival assay, and the measurement of bacterial numbers from cysts were performed, and nonpathogenic E. coli K12 was also applied. The association ratio of E. coli K1 with A. castellanii was 4.3 cfu per amoeba for 1 hr but E. coli K5 with A. castellanii was 1 cfu per amoeba for 1 hr. By invasion and survival assays, E. coli K5 was recovered less than E. coli K1 but still alive inside A. castellanii. E. coli K1 and K5 survived and multiplied intracellularly in A. castellanii. The survival assay was performed under a favourable condition for 22 hr and 43 hr with the encystment of A. castellanii. Under the favourable condition for the transformation of trophozoites into cysts, E. coli K5 multiplied significantly. Moreover, the pathogenic potential of E. coli K1 from A. castellanii cysts exhibited no changes as compared with E. coli K1 from A. castellanii trophozoites. E. coli K5 was multiplied in A. castellanii trophozoites and survived in A. castellanii cysts. Therefore, this study suggests that E. coli K5 can use A. castellanii as a reservoir host or a vector for the bacterial transmission.

• Journal of Plant Biotechnology
• /
• v.31 no.4
• /
• pp.255-259
• /
• 2004

Agrobacterium tumefaciens-mediated cotyledonary node transformation was used to produce transgenic soybean. Cotyledonary node explants of three cultivars and one genotype were co-cultivated with strains Agrobacterium (LBA4404, GV3101, EHA101, C58) containing the binary vectors (pCAMBIA3301 and pPTN289) carrying with CaMV 35S promoter-GUS gene as reporter gene and NOS promoter-bar gene conferring resistance to glufosinate (herbicide Basta) as selectable marker. There was a significant difference in the transformation frequency depend on bacteria strain. The EHA101 strain of the bacterial strains employed gave the maximum efficiency (3.6%). One hundred-six lines transformed showed the resistance in glufosinate. Histochemical GUS assay showed that at least 11 plants transformed with the GUS gene were positive response. The soybean transformants were obtained from the Thorne (5 plants), 1049 (5 plants) and Bakun (1 plant), respectively. Southern blot analysis and leaf painting assay revealed that the GUS and bar gene segregated and expressed in their progeny.

• Mycobiology
• /
• v.49 no.5
• /
• pp.491-497
• /
• 2021

An endolichenic fungus Xylaria grammica EL000614 produces grammicin, a potent nematicidal pyrone derivative that can serve as a new control option for root-knot nematodes. We optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for X. grammica to support genetic studies. Transformants were successfully generated after co-cultivation of homogenized young mycelia of X. grammica with A. tumefaciens strain AGL-1 carrying a binary vector that contains the bacterial hygromycin B phosphotransferase (hph) gene and the eGFP gene in T-DNA. The resulting transformants were mitotically stable, and PCR analysis showed the integratin of both genes in the genome of transformants. Expression of eGFP was confirmed via fluorescence microscopy. Southern analysis showed that 131 (78.9%) out of 166 transformants contained a single T-DNA insertion. Crucial factors for producing predominantly single T-DNA transformants include 48 h of co-cultivation, pretreatment of A. tumefaciens cells with acetosyringone before co-cultivation, and using freshly prepared mycelia. The established ATMT protocol offers an efficient tool for random insertional mutagenesis and gene transfer in studying the biology and ecology of X. grammica.