• Title/Summary/Keyword: Gel electrophoresis

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Effect of heat treatment on physicochemical properties of soybean (열처리 방법에 따른 대두의 이화학적 특성 변화)

  • Kim, Sun Hee;Jung, Eun Suk;Kim, So Young;Park, Shin Young;Cho, Yong Sik
    • Food Science and Preservation
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    • v.24 no.6
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    • pp.820-826
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    • 2017
  • Soybean is one of the most common food materials for making traditional Korean foods such as soybean paste, soy source and soy snack, and their manufacturing processes include heat treatment of soybean. This study was carried out to investigate the effect of heat treatment on the physicochemical properties of soybean. All samples were heat treated under commercial steamed, puffed or air-fried conditions, and then the protein molecular weight distribution, thermal properties, fluorescence intensity, protein solubility, and water and oil holding ability of the heat treated soybeans were examined. Sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated that heat treatment caused fragmentation of polypeptide chain in soybean, showing the band of low molecular ranging from 17 to 40 kDa. The differential scanning calorimetric analysis showed the decrease of enthalpy values (${\Delta}H$) by heat treatment. Fluorescence spectroscopy indicated that the heat treatment caused lipid oxidation as proved by increasing emission intensity. The protein solubility at pH 3-6, and water holding capacity of heat treated soybeans were the higher than no treatment. These results suggest that the heat treatment resulted in decreased enthalpy values, and increased protein degradation, lipid oxidation and water affinity of soybean. Moreover, the effect of heat treatment on physiochemical properties of soybeans was more significant under air-fried condition.

Optimized Condition of Genomic DNA Extraction and PCR Methods for GMO Detection in Potato (유전자재조합 감자의 검정을 위한 DNA분리 및 PCR검출의 최적조건 탐색)

  • Shin, Weon-Sun;Kim, Myung-Hee
    • Korean Journal of Food Science and Technology
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    • v.35 no.4
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    • pp.591-597
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    • 2003
  • To compare the quality of genomic DNA extracted from potato for PCR detection, four different methods, such as silica-based membrane method, silica-coated bead method, STE solution treatment, and CTAB-phenol/chloroform method, were evaluated. Also, to remove an excessive carbohydrate from the potato, ${\alpha}$- and ${\beta}$-amylase were used individually and in combination. When used both silica-based membrane method and silica-coated bead method combined with enzymes, the genomic DNAs were extracted from the raw potato with high purity for PCR. However, the silica-coated head method combined with enzyme treatment was the most efficient for extraction of the genomic DNA from the frozen fried potatoes. When applied with STE solution, the highly purified DNA was extracted from the raw potatoes without enzyme treatment in adequate yield for PCR. In cases of processed potatoes, such as frozen-fried potato and fabricated potato chips, CTAB-phenol/chloroform method is mostly feasible for DNA extraction and PCR efficacy at high sensitivity. As the results of PCR amplification, 216bp of PCR product was detected on 2% agarose gel electrophoresis, but any amplicons derived from New leaf and New leaf Y gene was not detected in any sample.

Partial Purification and Quantification of Insulin-like Growth Factor-I from Red Deer Antler (녹용으로부터 Insulin-like Growth Factor-I의 일부정제 및 정량)

  • Gu, Lijuan;Mo, Eun-Kyoung;Fang, ZheMing;Sun, BaiShen;Zhu, XueMei;Sung, Chang-Keun
    • Journal of Life Science
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    • v.17 no.10
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    • pp.1321-1329
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    • 2007
  • Deer antler tissue contains the most rapidly growing bone in the animal kingdom. Thus, it is likely that growing antler tissue is a rich source of local paracrine bone-stimulating factors. Growth factors, at least the insulin-like growth factor (IGF), control the bone-remodelling process. In this study, we tried to isolate and purify IGF-I from fresh antler tissue by the routine isolation and purification of protein. The purification involved ammonium sulfate precipitation, DEAE-Sepharose CL-60 ion-exchange chromatography, CM-Sepharose CL-6B ion-exchange chromatography, and Sephadex G-50 chromatography. Purified fractions from each step were analyzed by high-performance liquid chromatography (HPLC), SDS polyacrylamide gel electrophoresis (SDS-PACE), Dot-blot, and Western-blot methods. Furthermore, the quantification of partially purified IGF-I was calculated by enzyme-linked immunosorbent assays (ELISA) using antibody to human recombinant IGF-1. SDS-PAGE analysis of the final fraction yielded two molecular bands and the signal band was at 12 kDa on the Western-blot film. This purified IGF-I fraction showed a peak at retention time of eight min. The quantity of IGF-I in 20 g deer antler tissue as starting weight was calculated using a standard curve to be 2910 ng/ml, and total IGF-I amount is 0.291 g. The results show that IGF-I, which can be found in deer antler, can be partially purified and quantified by classic protein isolation methods.

Biological Activity and Biochemical Properties of Silkworm (Bombyx mori L.) Powder Fermented with Bacillus subtilis and Aspergillus kawachii (유용식용 균주에 의한 발효 누에분말의 이화학적 특성과 생리활성)

  • Cha, Jae-Young;Kim, Yong-Soon;Ahn, Hee-Young;Kang, Min-Jung;Heo, Su-Jin;Cho, Young-Su
    • Journal of Life Science
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    • v.21 no.1
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    • pp.81-88
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    • 2011
  • Biological activities (${\alpha},{\alpha}'$-diphenyl-${\beta}$-picrylhydrazyl (DPPH) free radical scavenging activity, fibrinolytic activity and reducing power) and biochemical properties (protein content and electrophoretical protein patterns) were examined in solid state fermentation with Bacillus subtilis and Aspergillus kawachii using silkworm powder (SP) as substrate. The highest protein contents and free radical scavenging activities were seen in the SP fermented for 12 days with B. subtilis and A. kawachii, and these were in a time-dependent manner. The highest reducing power was seen in the SP fermented for 6 days with B. subtilis and for 12 days with A. kawachii, respectively. The highest fibrinolytic activities were seen in silkworm fermented for 6 days with B. subtilis and A. kawachii, but this activity was higher in the A. kawachii fermented SP than that of B. subtilis. When total protein patterns were analyzed by SDS-polyacrylamide gel electrophoresis (PAGE), the proteins of the SP fermented with B. subtilis for 3 days were completely degraded, while the protein degradation in the SP fermented with A. kawachii occurred after 12 days and this degradation increased proportionally to culture time. As a result, the SP fermented with both B. subtilis and A. kawachii showed higher fibrinolytic activities after 6 days of fermentation and antioxidative activity after 12 days, indicating that physiological activities of the fermented SP using these strains were highly improved compared to the unfermented SP, and that this compound could be a candidate material as a dietary supplement of healthy functional foods.

Molecular Characterization of Clinically Isolated Staphylococcus aureus (인천지역에서 분리된 황색포도상구균의 분자생물학적 특성 분석)

  • Oh, Bo-Young;Kim, Jung-Hee;Gong, Young-Woo;Lee, Jae-Mann;Go, Jong-Myoung;Kim, Yong-Hee
    • Korean Journal of Microbiology
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    • v.44 no.4
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    • pp.305-310
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    • 2008
  • Staphylococcus aureus is one of the most significant pathogens and a causative agents of nosocomial infections. The emergence of methicillin resistant S. aureus (MRSA), in particular, has become a major clinical and epidemiological problems worldwide. In this study, we analyzed the toxin genes and investigated molecular epidemiological characteristics of S. aureus isolated from stools of diarrheal patients at the hospitals in Incheon. Of the 609 strains from 2,281 specimens, 173 strains retained enterotoxin; 68 isolates (39.30%), 100 isolates (57.80%) were classified to A and C type, respectively. In the antibiotic susceptibility, all of enterotoxin positive isolates were resistant to oxacillin. Eighty eight strains (50.86%) of 173 MRSA isolate possessed tsst gene, but eta and eth genes were not detected at all. In the detection of MRSA associated genes by PCR method, mecA genes were detected in 167 strains (96.53%). From the result of PFGE analysis, we classified tsst-positive MRSA to 10 types and 24 subtypes. Type A, H and F were the major strains comprised of 57.95% (51 strains), 10.22% (9 strains) and 9.09% (8 strains) respectively.

Fibrinolytic Activities and Effects of Gamma-Irradiated on Seeds from Coix lacryma-jobi L. Carthamus tinctorius L. and Malva verticillata L. (율무, 홍화, 아욱종자의 혈전용해 효소활성 및 감마선 조사의 영향)

  • Kwon Su-Jung;Lim Chae-Young;Kim Jae-Sung;Park Min-Hee;Lee Sook-Young
    • KSBB Journal
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    • v.21 no.1 s.96
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    • pp.20-27
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    • 2006
  • The fibrinolytic activities of soluble proteins extracted from seeds of Coix lacryma-jobi L., Carthamus tinctorius L. and Malva venicillata L. were studied. Fibrinolytic activity of extract from C. lacryma-jobi L. showed 1.3 times higher than plasmin used as positive control. The fibrinolytic enzyme was confirmed and extracted directly from seed of C. lacryma-jobi L. by a fibrin zymography. The protein was composed of a single polypeptide and its apparent molecular weight was found to be 7.8 kDa, as judged by SDS-polyacrylamide gel electrophoresis. The effect of temperature for the proteolytic enzyme activity were stabilized above $50^{\circ}C$ and then dramatically decreased. Also, the enzyme activity was clearly inhibited by APMSF, PMSF and TPCK, suggesting that it is a member of the chymotrypsin-like serine pretense. In addition, effects of gamma-irradiated on seed of each plants were revealed that 8 Gy and 64 Gy were higher than others. This result shown that gamma-irradiation of seeds were capable to increase the fibrinolytic activity. All these results suggest the pretense is a fibrinolytic enzyme belong to a family of chymotrypsin-like serine pretense.

Cloning and Expression of Thermostable $\beta$-Glycosidase Gene from Thermus filiformis Wai33 A1 in Escherichia coli and Enzyme Characterization

  • Kang, Sang-Kee;Cho, Kwang-Keun;Ahn, Jong-Kun;Kang, Seung-Ha;Han, Kyung-Ho;Lee, Hong-Gu;Choi, Yun-Jaie
    • Journal of Microbiology and Biotechnology
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    • v.14 no.3
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    • pp.584-592
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    • 2004
  • A thermostable $\beta$-glycosidase gene, tfi $\beta$-gly, was cloned from the genomic library of Thermus filiformis Wai33 A1. ifi $\beta$-gly consists of 1,296 bp nucleotide sequence and encodes a polypeptide of 431 amino acids. It shares a strong amino acid sequence similarity with the $\beta$-glycosidases from other Thermus spp. belonging to the glycosyl hydrolase family 1. In the present study, the enzyme was overexpressed in Escherichia coli BL21 (DE3) using the pET21b(+) vector system. The recombinant enzyme was purified to homogeneity by heat treatment and a $Ni^{2+}$-affinity chromatography. Polyacrylamide gel electrophoresis (PAGE) showed that the recombinant Tfi $\beta$-glycosidase was a monomeric form with molecular mass of 49 kDa. The temperature and pH range for optimal activity of the purified enzyme were 80- $90^{\circ}C$ and 5.0-6.0, respectively. Ninety-three percent of the enzyme activity was remained at $70^{\circ}C$ after 12 h, and its half-life at $80^{\circ}C$ was 6 h, indicating that Tfi $\beta$-glycosidase is highly thermostable. Based on its K_m$, or $K_{cat}K_m$, ratio, Tfi $\beta$-glycosidase appeared to have higher affinity for $\beta$-D-glucoside than for $\beta$-D-galactoside, however, $K_{cat} for \beta$-D-galactoside was much higher than that for $\beta$-D-glucoside. The activity for lactose hydrolysis was proportionally increased at $70^{\circ}C$ and pH 7.0 without substrate inhibition until reaching 250 mM lactose concentration. The specific activity of Tfi TEX>$\beta$-glycosidase on 138 mM lactose at $70{^\circ}C$ and pH 7.0 was 134.9 U/mg. Consequently, this newly cloned enzyme appears to have a valuable advantage of conducting biotechnological processes at elevated temperature during milk pasteurization in the production of low-lactose milk.

Studies on the Effects of Ozone Gas in Paddy Rice;4. Effects of Ozone Gas on Rice Growth at Different Nutrition Levels (수도생육(水稻生育)에 대(對)한 Ozone가스의 영향(影響)에 관(關)한 연구(硏究);4. 수도 (水稻)의 영양상태(營養狀態)와 Ozone가스 피해(被害))

  • Kim, Bok-Young;Lee, Suk-Hee;Kim, Maun-Soo
    • Korean Journal of Environmental Agriculture
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    • v.6 no.1
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    • pp.38-43
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    • 1987
  • This study was carried out to investigate the effect of ozone gas on paddy rice at the different nutrition levels. Jinjubyeo variety of rice plant was exposed to 0.3 ppm ozone gas for 3 hours. It was cultivated at three different application levels, optimum, -50%, and +50% of optimum of nitrogen, phosphorus, and potassium, respectively. After ozone gas fumigation, percentage of damaged leaf, malondialdehyde contents, activity of peroxidase, and nutrient contents of rice plant were observed. The results obtained are as follows. 1) Percentage of damaged leaf was increased at the both additional 50% application of nitrogen and 50% reduction of potassium. 2) Malondialdehyde contents of leaves were increased with the ozone gas exposure. 3) Percentage of damaged leaf was increased at the lower level of $K_2O/N$ ratio in leaves. 4) Nitrogen, phosphorus, and potassium contents in rice leaves were decreased with the ozone gas exposure. 5) The peroxidase bands on gel in electrophoresis were changed by the ozone gas exposure.

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Changes of Microbial Community Structure According to a Changes of Season and Influent Characteristics in Biological Wastewater Treatment (생물학적 폐수처리 공정에서의 계절 및 유입수 성상 변화에 따른 미생물 군집 특성 변화)

  • Son, Hyeng-Sik;Son, Hee-Jong;Kim, Mi-A;Ryu, Eun-Yeon;Lee, Geon;Lee, Sang-Joon
    • Journal of Korean Society of Environmental Engineers
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    • v.32 no.8
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    • pp.780-786
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    • 2010
  • The bacterial community structure in biological reactor in wastewater treatment system was investigated by denaturing gradient gel electrophoresis (DGGE) and fluorescent in situ hybridization (FISH). Samples were collected at different three points in wastewater treatment system. Through treatment processes, BOD (biochemical oxygen demand) and COD (chemical oxygen demand) of was removal efficiency was 83.1~98.6%, 67.2~85.2% respectively. Microbial community of aerobic tank and oxic tank were similar but anoxic tank was different (RRP group was increased about tripple) by DGGE and FISH in sludge (2007 October and 2008 January). Samples in 2007 October and 2008 January were dominant ${\alpha}$-Proteobacteria and CF group respectively. Sludge in 2008 April were different comparing former results dominant others as 65~80%. Others group was dominant. Eubacteria by FISH with the probe EUB338 was about $1.7{\sim}7.6{\times}10^9\;cells/mL$. It could be successfully observed bacterial community in biological wastewater system.

Effects of Transgenic Soybean Cultivation on Soil Microbial Community in the Rhizosphere (형질전환 콩 재배가 근권 토양 미생물상에 미치는 영향)

  • Lee, Ki-Jong;Sohn, Soo-In;Lee, Jang-Yong;Yi, Bu-Young;Oh, Sung-Dug;Kweon, Soon-Jong;Suh, Seok-Choel;Ryu, Tae-Hun;Kim, Kyung-Hwan;Park, Jong-Sug
    • Korean Journal of Environmental Agriculture
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    • v.30 no.4
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    • pp.466-472
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    • 2011
  • BACKGROUND: Soybean [Glycine max (L.) Merrill] is a legume and an important oil crop worldwide. This study was conducted to evaluate the possible impact of transgenic soybean cultivation on the soil microbial community. METHODS AND RESULTS: Microorganisms were isolated from the rhizosphere soils. Microbial community was identified based on the culture-dependent and molecular biology methods. The total numbers of bacteria, fungi, and actinomycete in the rhizosphere soils cultivated with transgenic and non-transgenic soybeans were similar to each other, and there was no significant difference between transgenic and non-transgenic soybeans. Dominant bacterial phyla in the rhizosphere soils cultivated with transgenic or non-transgenic soybeans were Actinobacteria, Firmicutes, and Proteobacteria. The microbial communities in transgenic and non-transgenic soybean soils were characterized using the denaturing gradient gel electrophoresis (DGGE). The DGGE profiles showed the different patterns, but didn't show significant difference to each other at 0.05 significance level. DNAs were isolated from soils cultivating transgenic or non-transgenic soybeans and analyzed for persistence of transgenes in the soil by using PCR. PCR analysis revealed that there were no amplified ${\gamma}$-tmt and bar gene in soil DNA. CONCLUSION(S): The results of this study suggested that microbial community of soybean field were not significantly affected by cultivation of the transgenic soybeans.