• Journal of Microbiology and Biotechnology
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• 제5권4호
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• pp.194-199
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• 1995

A portion (7.4 kb) of ribosomal DNA tandem repeat unit from a genome of the mushroom T. matsutake has been cloned. A 1.75 kb EcoRI fragment was cloned first using S. cerevisiae 255 rRNA gene as a probe, and this was then used for further cloning. A chromosomal walking experiment was carried out and the upstream region of the 1.75 kb fragment was cloned using SmaI/BamHI enzyme, the size was estimated to be 5.2 kb in length. Part of the downstream region of the 1.75 kb fragment was also cloned using XbaI/BamHI enzymes. Restriction enzyme maps of three cloned DNA fragments were constructed. Northern hybridization, using total RNA of T. matsutake, and the restriction fragments of three cloned DNAs as probes, revealed that all four ribosomal RNA genes (large subunit[LSU], small subunit [SSU], 5.85 and 5S rRNA genes) are present in the cloned region. The gene organization of the rDNA are regarded as an intergenic spacer [IGS]2 (partial) - SSU rRNA - internal transcribed spacer [ITS]1 - 5.8S rRNA - ITS2 - LSU rRNA - IGS1 -5S rRNA - IG52 (partial).

• Restorative Dentistry and Endodontics
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• 제20권2호
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• pp.577-609
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• 1995

Bacteria have been regarded as one of the most important factors in pulpal and periapical diseases. Streptococci are frequently isolated facultative anaerobes in infected root canals. Recently molecular biological techniques have been rapidly progressed. This study was designed to apply the molecular biological tools to the identification and classification of streptococci in the endodontic microbiology. Streptococci isolated from infected root canals were identified with both Vitek Systems and API 20 STREP. Identification results were somewhat different in several strains of streptococci. Eighteen streptococci and enterococcal was difficult so to digest plasmid DNA using Hind III and EcoRI to differentiate strains by restriction enzyme analysis of plasmid DNA. 16S rDNA of chromosome was amplified by polymerase chain reaction(PCR) and then restricition fragment length polymorphism(RFLP) using several restriction enzymes was observed. The molecular mass of 16S rDNA of chromosomal DNA was approximately 1.4kb. There were three to five RFLP patterns using eight restriction enzymes. RFLP patterns digested with CfoI which recognizes four base sequences were identical in all stains. Hind III which recognizes six base sequences could not digest the 16S rDNA. Restriction enzymes which recognize five base sequences were suitable for RFLP pattern analysis. At least three different restriction enzymes were needed to compare each strains. 16S rDNA PCR-RFLP was simple and rapid to differentiate and classify strains and could be used in the epidemiological study of root canal infections.

• Journal of Microbiology and Biotechnology
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• 제12권1호
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• pp.176-181
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• 2002

In order to isolate and identify Bifidobacterium spp. that originated in Korea, feces were sampled from healthy Korean adults and children living in three villages, the first having a history of longevity and the other two where the diet did not include fermented milk or any pharmaceutical preparations. Through the use of Gram staining and microscopic examination for cell morphology, 23 bacterial strains presumed to be the Bifidobacterium genus were isolated from the feces of 13 out of a total of 59 Korean people. To identify the Bifidobacterium strains at the genus level, these bacteria were then analyzed by TLC and the fructose-6-phosphate phosphoketolase (F6PPK) test. The result showed that 22 of the isolated strains were confirmed to be members of the genus Bifidobacterium. All of these bifidobacteria were also identified as Bifidobacterium spp. by the fermentation test. Using a RFLP analysis, an attempt was made to identify the Bifidobacterium spp. that had been isolated from both Korean adults and children. In a genomic Southern blot analysis after digestion with two restriction enzymes (EcoRI, HindIII), all of the 14 randomly selected Korean isolates showed patterns identical to those of three different B. longum species. Another restriction enzyme, CfoI (4-bp recognition enzyme), was then used to identify the strain. Interestingly, all the Korean isolates were identified as B. longum ATCC 15708, indicating that a RFLP analysis was effective for identifying Bifidobacterium spp. at both the strain and species levels.

• 한국미생물·생명공학회지
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• 제20권3호
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• pp.289-294
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• 1992

토양에서 분리한 여러 알칼리성 세균을 대상으로 제한효소의 유무를 조사하여 Bacillu sp. 8-13으로부터 제한효소의 활성을 발견할 수 있었다. 이 균주의 제한효소는 streptomycin sulfate 침전, ammonium sulfate 침전, DEAD-cellulose와 phosphocellulose ion exchange colum chromatography와 phosphocellulose ion exchange colum chromatography 과정을 통하여 정제하였고 0.1 SDS를 포함하는 7.5 polyacrylamide gel electrophoresis로 subunit의 분잘량을 조사한 결과 약 32,000 dalton이었다. 또 특히 이 균주는 형태적 배양학적 특성이 Bacillus alkalophilus subsp. haldurans와 매우 유사하며, 최적 생육, pH는 10.3이었고, 최적 증식온도는 $50^{\circ}C$ 이었다.

• BMB Reports
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• 제39권2호
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• pp.140-144
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• 2006

A deoxyriboendonuclease has been purified to near homogeneity from a fast growing mycobacterium species, M. smegmatis and characterized to some extent. The size of enzyme is about 43 kDa as determined by a denaturing gel analysis. It shows optimum activity at $32^{\circ}C$ in Tris-HCl buffer (pH 7.2) containing 2.5 mM of $MgCl_2$. Both EDTA and $K^+$ but not $Na^+$ inhibit its activity. Evidences show that the enzyme is not a restriction endonuclease but catalyzes the endonucleolytic cleavage of both the double- as well as the single-strand DNA non-specifically. It has been shown that the cleavage by this enzyme generates DNA fragments carrying phosphate groups at 5' ends and hydroxyl group at the 3' ends, respectively. Analysis reveals that no endonuclease having size and property identical to our deoxyriboendonuclease had been purified from M. smegmatis before. The property of our enzymes closely matches with the deoxyriboendonucleases purified from diverse sources including bacteria.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 1986년도 추계학술대회
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• pp.509.1-509
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• 1986

Recent advances in recombinant DNA techniques have provided a tool for breeding of microorganisms of hyper production. Enzyme production by cloned microorganism has some advantages. They are ⅰ) Enzymes can be produced by a microorganism easily cultured ⅱ) Hyper production. ⅲ) In some cases, such as thermophilic enzyme gene is cloned in a mesophilic bacteria, the enzyme purification procedure can be simplified. One example, production of thermophilic ${\beta}$-galactosidase in B. subtilis will be presented. Bacillus stearothermophilus IAM 11001 produced three ${\beta}$-galactosidases, ${\beta}$-galactosidase I, II and III (${\beta}$-gal-I, II and III). By connecting restriction fragments of the chromosomal DNA to plasmid vector, followed by transformation of Escherichia coli, two ${\beta}$-galactosidase genes (bgaA and bgaB) located close to each other on the chromosome were cloned.

• 미생물학회지
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• 제27권3호
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• pp.188-193
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• 1989

Gene for lactose catabolism in Lactobacillus casei SW-M1 was encoded by a 60Kb metabolic plasmid. A derivative of only 10kb, pPlac 15 of recombinant plasmid, was constructed by introducing into pBR322 and was cloned into E. coli using restriction endonuclease Pst I. A 10kb insery DNA in plasmid pBR322 was identified as a gene encoded phospho-$\beta$-galactosidase by the determination of enzyme activity. Phospho-$\beta$-galactosidase was apparently expressed in E. coli. The enzyme activities of cell-free extract from transformant E. coli HB101 carrying pPLac 15 DNA were not different from that of L. casei as a donor strain on the basis of enzyme properites. However, specific activity of phospho-$\beta$-galactosidase in the cloned strain with Lac $Y^{-}$ phenotype of E. coli HB101 was lower than that in L. casei strain.

• Journal of Nutrition and Health
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• 제6권3호
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• pp.1-8
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• 1973

Metabolic responses to the protein-free, high-carbohydrate diet and subsequent food restriction on the same diet at the level of 50% and 75% has been studied on the adult albino rats. The energy source was either corn starch or sugar. In experiment I, adult male rats weighing $509{\pm}8g$ were divided into two groups 10 rats each. Rats fed on the stock diet served as a control. Rats of restriction group received a protein free diet until they reduced their weight down to 400g and continue on a protein-calorie restriction diet until they reduced their weight down to 300g. In experiment II, 28 adult male rats and the same numbers of female rats weighing $329{\pm}5g$ and $223{\pm}4g$ respectively were divided into four groups, 7 males and females in each. Rats fed on a stock diet were sacrificed at the point when others started a protein free diet. These were served as the control. The protein free group received a protein free diet ad libitum for 4 weeks. The 50% restriction group and 75% restriction group were fed on a protein free diet coupled with food restriction at levels of 50% and 75% respectively for 3 weeks. In the result of this study: 1. The rate of body weight changes was similar between the males and the females. Feeding protein free diet ad lib. initiated a rapid weight lost of approximately 25% and protein free diet coupled with food restriction showed 37-43% reduction of their initial weight. 2. There was no significant differences in the value of the N concentration in liver, spleen, brain and muscle between controls and experimental groups. 3. Rats fed on protein free diet showed 1/10 value of the control in the nitrogen excretion in urine. However female showed less N excretion than male. 4. Observing blood picture, the effects of protein depletion and calorie restriction were not appeared any remarkable changes. 5. There was no sign of fatty liver which might result from protein depletion and calorie restriction. 6. Following semi-starvation, FAO and HMP-DH total enzyme activity was reduced, but activity per unit weight was relatively stable.

• 미생물학회지
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• 제23권2호
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• pp.115-121
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• 1985

Lactobacillus casei에 감염하는 독성 phage중 각 분류꾼을 대표하는 5종의 phage (J1, TK93, K1, PD 5 빛 CP 1)와 l 종의 용원 phage (${\phi}$ 1043) 의 핵산의 특성을 바교 검토하였다. 실험한 6 종의 phage는 모두 double S stranded DNA를 갖고 있였으며 J1. TK93, K1 및 ${\phi}$ 1043 DNA의 크기는 약 42Kb, PD5 와 CP1 DNA는 140K Kb정도로 서로 비슷하였다. EcoR 1으로 절단시 J1, TK93, K1, PD5, CP1 및 ${\phi}$1043은 각각 13, 13. 11, 14, 14와 12개의 절편을 갖는 특징적인 절단양식을 보여주었다. J1, TK93 및 ${\phi}$ 1043 DNA에는 cohesive end가 존재 하였고 K1, PD5 빛 CP1 DNA에는 없는 것으로 사료되었다. J1과 TK93 DNA의 제한효소 지도를 작성하여 비교 검토하였으며 이상의 결과로부터 진화적 유연관계를 검토하였다.

• Asian-Australasian Journal of Animal Sciences
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• 제24권6호
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• pp.752-756
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• 2011

Polymorphism of the second exon of the caprine leukocyte antigen-DRB3 gene (CLA-$DRB3^*02$) was investigated in this study. The 285 bp PCR product of 258 individuals from 10 domestic goat breeds in Southwest China was digested with restriction endonucleases PstI and HaeIII and then genotyped. Three alleles and 4 restriction digestion profiles were distinguished by digestion of the PCR fragment by PstI, and 8 alleles and 13 genotypes by HaeIII. For HaeIII restriction enzyme sites, the Chi-square ($X^2$) test showed that all goat breeds in this study did not fit with the Hardy-Weinberg equilibrium (p<0.01 or p<0.05). The highly polymorphic nature of CLA-$DRB3^*02$ was demonstrated and the ranges of gene heterozygosity (He) and polymorphism information content (PIC) were 0.36-0.63 and 0.32-0.55, respectively. Clustering analysis showed that the 10 goat breeds clustered into two groups and Dazu Black goat had a close genetic relationship with Chengdu Grey, Jintang Black and Nanjiang Yellow goats.