• Tuberculosis and Respiratory Diseases
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• v.49 no.5
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• pp.546-557
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• 2000

Background : Oligonucleotide chip technology has proven to be a very useful tool in the rapid diagnosis of infectious disease. Rifampin resistance is considered as a useful marker of multidrug-resistance in tuberculosis. Mutations in the rpoB gene coding $\beta$ subunit of RNA polymerase represent the main mechanism of rifampin resistance. The purpose of this study was to develop a diagnosis kit using oligonucleotide chip for the rapid and accurate detection of rifampin-resistance in Mycobacterium tuberculosis. Method : The sequence specific probes for mutations in the rpoB gene were designed and spotted onto the glass slide, oligonucleotide chip. 38 clinical isolates of Mycobacterium were tested. A part of rpoB was amplified, labelled, and hybridized on the oligonucleotide chip with probes. Results were analyzed with a laser scanner. Direct sequencing was done to verify the results. Result : The low-density oligonucleotide chip design어 to determine the specific mutations in the rpoB gene of M. tuberculosis accurately detected rifampin resistance associated with mutations in 28 clinical isolates. Mutations at codons 531, 526, and 513 were confirmed by direct sequencing analysis. Conclusion : Mutant detection using oligonucleotide chip technology is a reliable and useful diagnostic tool for the detection of multidrug-resistance in M. tuberculosis.

• Journal of Mushroom
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• v.7 no.3
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• pp.130-134
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• 2009

To develop new white variety of Oyster mushroom, all white varieties which have been collected and kept in the lab were revived and screened their cultural characteristics. 84 intra-specific Oyster mushroom hybrids between the white-colored mutants Suhan and Wonhyeong were developed using hyphal anastomosis technique in 2007. The Po2007-63 ($2842-7{\times}0205-7$) was shown the best cultural characteristics, selected to be a new variety and named as 'Goni'. The new commercial strain, 'Goni' has white pilei and grows well under spring and autumn conditions in Korea. The fruiting bodies of 'Goni' are of an excellent quality in that not only the stipe is thick and long but also the pileus is small and hard. The optimum temperatures for mycelial growth and fruiting body development were $25-30^{\circ}C$ and $10-16^{\circ}C$, respectively. Time period required for the initiation of the first fruiting body is about 3 to 5 days depending on the temperatures. The shape of fruiting body is thin funnel shape. Fruiting body production per bottle was about $91{\pm}13$ g which is almost 97% quantity compared to that of other variety 'Miso'. Relatively low temperature incubation ($11^{\circ}C$) resulted in the development of better quality of 'Goni' mushrooms. When two different media including potato dextrose medium and mushroom complete medium were compared, the growth of mushroom were much faster in mushroom complete medium at $20-25^{\circ}C$, but not at $25^{\circ}C$. Similar results were observed with other variety 'Miso'. Analysis of the genetic characteristics of the new commercial strain 'Goni' showed a major DNA profile as that of the parental Suhan when primer URP 1 was used, but different to 'Miso' that was used as a control. When screens were performed with primer URP 2, DNA patterns were similar both to that of the parents and 'Miso'. This new variety of the white Oyster mushroom has a clean and fresh image that corresponds well to "health food". We therefore expect that this new strain will satisfy the consumers demand for variety and excellent mushrooms.

• Journal of Mushroom
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• v.11 no.3
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• pp.154-158
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• 2013

To develop a new variety of oyster mushroom(Pleurotus ostreatus), parental strains was selected by the method of Mon-Mon crossing between monokaryotic strains derived from ASI 2596(Suhan No.3) and ASI 2782(Black pileus mutant). The SB-73(ASI 2596-11 x 2782-8) was shown the best cultural characteristics, selected to be a new variety and named as 'Dagul'. The 'Dagul' was formed incompatibility line distinctly in the confrontation growth of parental strains Suhan No.3 and ASI 2782. The optimum temperature for mycelial growth, fruiting body development and pH arrange were $25{\sim}30^{\circ}C$, $14{\sim}17^{\circ}C$ and pH5~8, respectively. Fruiting body production per bottle was about $68.0{\pm}24.1$ g which is almost 115% quantity compared to that of other variety Suhan No.3. And also the stipe is long and individual generation is multiple. Analysis of the genetic characteristics of the new variety 'Dagul' showed different DNA bands as that of the control strains, Suhan No.3 and ASI 2782, when RAPD(Random Amplified Polymorphic DNA) primers URP7 and Rcb1 were used. This new variety 'Dagul' of oyster mushroom is characterized by multiple of individual generation and the stipe is long. We therefore expect that this new strain will increase of the income by cultivation of field.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.3
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• pp.183-190
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• 2004

Objectives: Methylenetetrahydrofolate reductase (MTHFR) mutation are commonly associated with hyperhomocysteinemia, and through their defects in homocysteine metabolism, they have been implicated as a risk factor for recurrent spontaneous abortion. Recent report describe that 28-bp tandem repeat polymorphism in thymidylate synthase enhancer region (TSER) that influence enzyme activity would affect plasma homocysteine level. We have investigated the relationship between TSER genotype and plasma homocysteine level in 54 patients with recurrent spontaneous abortion. Methods: Plasma homocysteine level was measured by fluorescent polarizing immunoassay. MTHFR mutation (C677T and A1298C) was identified by PCR-restriction fragment length polymorphism assay and TSER mutation was analyzed by PCR method. The data were analyzed using the program SAS 8.2 for Windows. Results: Total homocysteine level was significantly higher in MTHFR 677TT genotype ($9.80{\pm}3.87{\mu}mol/L$) than MTHFR 677CC genotype ($8.14{\pm}1.74{\mu}mol/L$) in Korean patients with unexplained recurrent spontaneous abortion (p=0.0143). However, the plasma homocysteine level was not significantly different in the MTHFR 1298AA ($8.42{\pm}2.65{\mu}mol/L$) and 1298CC ($6.09{\pm}0.32{\mu}mol/L$; p=0.2058) and, TSER 2R2R ($8.61{\pm}1.68{\mu}mol/L$) and 3R3R ($8.05{\pm}2.81{\mu}mol/L$; p=0.9319) mutant genotypes, respectively. In this study, we found the combination effects of TSER and MTHFR C677T genotypes. Plasma homocysteine levels were the highest ($11.47{\pm}4.66{\mu}mol/L$) in individuals with TSER 3R3R ($8.05{\pm}2.81{\mu}mol/L$) and MTHFR 677TT ($9.80{\pm}3.87{\mu}mol/L$) genotypes. Individuals with a combination of both TSER 2R2R/2R3R and MTHFR 677CC/CT genotypes ($7.69{\pm}1.77{\mu}mol/L$) had lower plasma homocysteine levels than TSER 2R2R ($8.61{\pm}1.68{\mu}mol/L$) and MTHR 677CC ($8.14{\pm}1.74{\mu}mol/L$) genotypes, respectively. The effect of MTHFR polymorphism in the homocysteine metabolism appears to be stronger than that of TSER polymorphism. Conclusion: Although statistically not significant, we found the elevated level of plasma homocysteine in combined genotypes with TSER and MTHFR (C677T and A1298C) in Korean patients with unexplained habitual abortion. In this study, we reported the possibility that TSER polymorphism is a genetic determinant of plasma homocysteine levels in the Korean patients as well as MTHFR C677T polymorphism. A large prospective study is needed to verify our findings.

• Journal of The Korean Society of Grassland and Forage Science
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• v.24 no.1
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• pp.61-70
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• 2004

Silage additives are needed to increase the quality of whole crop rice silage which seldom produce without the additives due to both high pH and butyric acid concentrations. Little information, however, is available about the silage fermentation of whole crop rice added with silage additives in Korea. This study was conducted to determine the optimum levels of silage additives by evaluating the effects of latic acid bacteria (LAB) and formic acid concentrations on the silage quality of whole crop rice harvested at different mature stages. Field study was established early in May until October 7th on a rice field at Yupori, Sinbuk-yeup, Chunchon, Kangwon-Do. "Ilpum" mutant rice was harvested at six different mature stages; booting stage (17 Aug.), milk-ripe stage (27 Aug.), dough stage (7 Sep.), yellow ripe stage (17 Sep.), dead ripe stage (27 Sep.) and full ripe stage (7 Oct.). Each sample was ensiled in three different ways; with 1) LAB (0.05, 0.1 and 0.2% of sample wt), 2) formic acids (0.2, 0.3 and 0.4% of sample wt.) and 3) no additive. The additive levels did not affect dry matter content, crude protein, fiber and total digestable neutriant concentrations at all stages. Addition of additives significantly decreased the silage pH and butyric acid concentrations which tended to be more decreased with higher levels of additives. Latic acid concentrations were higher with the use of additives, especially with LAB. The lower concentrations of ammonia-N were observed in additive treatments at all stages, but the concentrations of ammonia-N did not differ according to the additve levels after yellow ripe stage (0.69, 0.60 and 0.71% of DM in 0.05, 01 and 0.2% of LAB, respectively; 0.64 0.59 and 0.75% of DM in 0.2, 0.3 and 0.4% of formic acid, respectively). These results indicate that the optimum addition levels of LAB and formic acid are 0.5∼0.1% and 0.2∼0.3%, respectively, on which the high quality of rice whole crop silage was produced. produced.

• Journal of Life Science
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• v.16 no.7 s.80
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• pp.1133-1140
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• 2006

Human HtrA1 (High temperature requirement protein A1) is a homologue of the E. coli periplasmic serine protease HtrA. A recent study has demonstrated that HtrA1 is a serine protease involved in processing of insulin like growth factor binding protein (ICFBP), indicating that it serves as an important regulator of IGF activity. Additionally, several lines of evidence suggest a striking correlation between proteolytic activity of HtrA1 serine protease and the pathogenesis of several diseases; however, physiological roles of HtrA1 remain to be elucidated. We used the pGEX bacterial expression system to develop a simple and rapid method for purifying HtrA1, and the recombinant HtrA1 protein was utilized to investigate the optimal conditions in executing its proteolytic activity. The proteolytically active HtrA1 was purified to approximately 85% purity, although the yield of the recombinant HtrA1 protein was slightly low $460{\mu}g$ for 1 liter E. coli culture). Using in vitro endoproteolytic cleavage assay, we identified that the HtrA1 serine protease activity was dependent on the enzyme concentration and the incubation time and that the best reaction temperature was $42^{\circ}C$ instead of $37^{\circ}C$. We arbitrary defined one unit of proteolytic activity of the HtrA1 serine protease as 200nM of HtrA1 that cleaves half of $5{\mu}M\;of\;{\beta}-casein$ during 3 hr incubation at $37^{\circ}C$. Our study provides a method for generating useful reagents to investigate the molecular mechanisms by which HtrA1 serine protease activity contributes in regulating its physiological function and to identify natural substrates of HtrA1.

• Journal of Life Science
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• v.22 no.12
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• pp.1628-1636
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• 2012

This study examined the antioxidant activity of the dark purple rice seeds from the rice line, MGI079, derived from insertional mutagenesis. The contents of polyphenolic compounds were 1.3 and 1.9-fold higher in the MGI079-2-1 and MGI079-2-6 rice lines than in the donor cultivar MGI079. Flavonoid contents were 6.4-fold higher in the MGI079-2-1 line. The MGI079-2-1 line showed a 24.4-fold higher activity in DPPH free radical scavenging compared to the MGI079 line. The anthocyanin content of the MGI079-2-6 line was more than 106.4-fold higher than the MGI079 line and 1.4-fold higher than the Heugnam line. Anthocyanin content in colored rice grains was negatively correlated with Hunter's L, a, and b values, with the correlation coefficients of $-5.64^{**}$, $5.21^{**}$ and -1.15, respectively. The grain length/width of a mutant of MGI079 segregated to a medium and bold type compared to the medium type of MGI079. However, the 1,000 grain weight was decreased to 13.6~19.6 g compared to 19.8 g for MGI079. Amylose content of the endosperm was 5.6~23.8% higher than in the MGI079 line. The grain of mutants of MGI079 was distinguished by its starch characteristics. The higher antioxidant activity of the MGI079-2-1 and MGI079-2-6 lines indicated functional characteristics associated with high-value resources, so future breeding should focus on the development of pigments in colored rice in new varieties.

• KSBB Journal
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• v.21 no.3
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• pp.204-211
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• 2006

For the production of the recombinant human interferon-gamma(rhIFN-${\gamma}$) in Escherichia coli, human glucagon and ferritin heavy chain were used as fusion partners. Even though rhIFN-${\gamma}$ is expressed as an inclusion body form in E. coli because of strong hydrophobicity of itself, over 50% of fused rhIFN-${\gamma}$ was expressed as soluble form in E. coli $Origami^{TM}$(DE3) harboring pT7FH(HE)-IFN-${\gamma}$ which encodes ferritin heavy chain-fused rhIFN-${\gamma}$. In the case of using glucagon-ferritin heavy chain hybrid mutant as a fusion partner, 6X His-tag was additionally introduced to N-terminus of GFHM(HE)-IFN-${\gamma}$ for enhancing purification yields of rhIFN-${\gamma}$. Fusion protein HGFHM(HE)-IFN-${\gamma}$ with two 6X His-tag was more effectively bound to Ni-NTA agarose bead than GFHM(HE)-IFN-${\gamma}$ with a 6X His-tag. rhIFN-${\gamma}$ was completely purified from enterokinase-treated HGFHM(HE)-IFN-${\gamma}$ by Ni-NTA affinity column. For high-level production of rhIFN-${\gamma}$, glucose was used as the sole carbon source with simple exponential feeding rate($2.4{\sim}7.2g/h$) in fed-batch process. The effective lactose concentration for the expression of the rhIFN-${\gamma}$ was $10{\sim}20mM$. Under the fed-batch culture conditions, rhIFN-${\gamma}$ production yield reached 11 g DCW/L for 6 hours after lactose induction.

• Tuberculosis and Respiratory Diseases
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• v.63 no.1
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• pp.31-41
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• 2007

Background: In pathogenesis and prognosis of lung cancer, significance of enormous types of genetic expression were very compounding and undetermined. We performed this study to search association between clinical characteristics and expression of COX-2, MMP-9 and p53 in non-small cell lung cancer. Methods: Ninety-one patients with adenocarcinoma or squamous cell carcinoma were enrolled. We had searched clinical data retrospectively and performed immunohistochemical staining for COX-2, MMP-9 and p53. We had analyzed significance of these three genes in clinical features and prognosis for survival. Results: 1) In squamous cell carcinoma, male was predominant and was significantly correlated with smoking. 2) Major prognostic determinants for overall survival were curative resection. 3) Expression of COX-2 was more frequent in adenocarcinoma than in squamous cell carcinoma. 4) Negative staining of COX-2, MMP-9 and p53 was more frequent in squamous cell carcinoma than adenocarcinoma. 5) Survival duration was longer in the group with positive expression of p53 and negative for COX-2 and MMP-9 (median duration of survival = 165.6 weeks) than groups with the other expressional patterns. 6) Significant correlation was found between expression of MMP-9 and COX-2. In squamous cell carcinoma, expression of MMP-9, COX-2 and mutant p53 were mutually correlated. 7) COX-2 expression was significant prognostic factor for survival in resected cancer group. In unresected inoperable non-small cell lung cancer group, MMP-9 was statistically significant prognostic factor for overall survival. Conclusion: COX-2 and MMP-9 might have some roles for progression or prognosis in some selected patients with non-small cell lung cancer. COX-2 and MMP-9 may have some roles for disease progression or prognosis in selected patients with NSCLC.

• Journal of Life Science
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• v.16 no.1
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• pp.37-43
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• 2006

Gap junction is an ion channel forming between adjacent cells. It also acts as a membrane channel like sodium or potassium channels in a single cell. The amino acid residues up to the $10^{th}$ position in the amino (N)-terminus of gap junction hemichannel affect gating polarity as well as current-voltage (I-V) relation. While wild-type Cx32 channel shows negative gating polarity and inwardly rectifying I-V relation, T8D channel in which threonine residue at $8^{th}$ position is replaced with negatively charged aspartate residue shows reverse gating polarity and linear I-V relation. It is still unclear whether these changes are resulted from the charge effect or the conformational change of the N-terminus. To clarify this issue, we made a mutant channel harboring cysteine residue at the $8^{th}$ position (T8C) and characterized its biophysical properties using substituted-cysteine accessibility method (SCAM). T8C channel shows negative gating polarity and inwardly rectifying I-V relation as wild-type channel does. This result indicates that the substitution of cysteine residue dose not perturb the original conformation of wild-type channel. To elucidate the charge effect two types of methaenthiosulfonate (MTS) reagents (negatively charged $MTSES^-$ and positively charged $MTSET^+$) were used. When $MTSES^-$ was applied, T8C channel behaved as T8D channel, showing positive gating polarity and linear I-V relation. This result indicates that the addition of a negative charge changes the biophysical properties of T8C channel. However, positively charged $MTSET^+$ maintained the main features of T8C channel as expected. It is likely that the addition of a charge by small MTS reagents does not distort the conformation of the N-terminus. Therefore, the opposite effects of $MTSES^-$ and $MTSETT^+$ on T8C channel suggest that the addition of a charge itself rather than the conformational change of the N-terminus changes gating polarity and I-V relation. Furthermore, the accessibility of MTS reagents to amino acid residues at the $8^{th}$ position supports the idea that the N-terminus of gap junction channel forms or lies in the aqueous pore.