• Title/Summary/Keyword: total RNA

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Isolation and Characterization of Pre-$tRNA^{Val}$ Splicing Mutants of Schizosaccharomyces pombe

  • Hwang, Ku-Chan;Kim, Dae-Myung
    • Journal of Microbiology
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    • v.35 no.4
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    • pp.334-340
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    • 1997
  • A collection of 132 temperature sensitive (ts) mutants was generated by the chemical mutagenesis of Schizosaccharomyces pombe wild type strain and screened for tRNA splicing defects on Northern blots by hybridization with an oligonucleotide that recognizes the exon of the S. pombe tRNA^Val as a probe. We identidied 6 mutants which accumulate precursor $tRNA^{Val}$. Among them, 2 mutants exhibited remarkable morphological differences compared to wild type cells. One tRNA splicing mutant showed elongated cell shape in permissive as well as non-permissive cultures. The other mutant exhibited shortened cell morphology only in nonpermissive culture. The total RNA pattern in the splicing mutants appeared to be normal. Genetic analysis of four $tRNA^{Val}$ splicing mutants demonstrated that the mutation reside in different genes.

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MicroRNA Expression in Leukemia Cell Line(K562 cell) Using Rhus Verniciflua Stokes (건칠(乾漆)을 이용한 K562 만성 골수성 백혈병 세포주에서의 MicroRNA 발현 규명)

  • Choi, Hyun Sook
    • The Korea Journal of Herbology
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    • v.34 no.6
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    • pp.71-78
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    • 2019
  • Objective : The purpose of the study was to identify expression profiling of miRNAs associated with cancers after treating allergen-removed Rhus Verniciflua Stokes and allergen-removed Rhus Verniciflua Stokes fumigaed Angelica gigas on leukemia cell lines. Methods : miRNA expression has been analyzed using miRNA array method through denaturation and hybridization after isolating the total RNA from leukemic cell line treated with 100 ㎍/㎖ of aRVS and aRVS-A each. Microarray expressions were interpreted as 'significant' on miRNAs when decreased less than 0.5 fold or increased more than 1.5 fold compared with the control group. Results : Among 158 miRNAs in total, 32 miRNAs were significantly presented in miRNAs expression. miRNA has been activated with a variety of genes for predicted targets, and the overexpressed miRNAs were categorized according to proliferation and metastasis of cancer in this study. The findings were reported that seven miRNAs (let-7b, miR-193a-5p, 296-3p, 26a, 22, 124a, 92b) showed significant expressions on proliferation and growth, seven miRNAs (miR-193a-5p, 26a, 200c, 183, 124a, 198, 210) presented meaningful expressions on invasion and metastasis, two miRNAs (let-7b, miR-210) were highly expressed on angiogenesis, five miRNAs (let-7b, miR-26a, 181d, 181c, 296-5p) related with apoptosis, and six miRNAs (let-7b, miR-200c, 183, 370, 124a, 191) were associated with prognosis of cancer and early diagnostic factors for cancer. Conclusion : The mechanism of miRNA takes a role in diagnosis, treatment, and prognotic factors for cancer as well. This study suggested that further detailed research on overexpression of specific miRNA should be carried out continuously in the future.

Comparison of Total RNA Isolation Methods for Analysis of Immune-Related microRNAs in Market Milks

  • Oh, Sangnam;Park, Mi Ri;Son, Seok Jun;Kim, Younghoon
    • Food Science of Animal Resources
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    • v.35 no.4
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    • pp.459-465
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    • 2015
  • Bovine milk provides essential nutrients, including immunologically important molecules, as the primary source of nutrition to newborns. Recent studies showed that RNAs from bovine milk contain immune-related microRNAs (miRNA) that regulate various immune systems. To evaluate the biological and immunological activity of miRNAs from milk products, isolation methods need to be established. Six methods for extracting total RNAs from bovine colostrums were adopted to evaluate the isolating efficiency and expression of miRNAs. Total RNA from milk was presented in formulation of small RNAs, rather than ribosomal RNAs. Column-combined phenol isolating methods showed high recovery of total RNAs, especially the commercial columns for biofluid samples, which demonstrated outstanding efficiency for recovering miRNAs. We also evaluated the quantity of five immune-related miRNAs (miR-93, miR-106a, miR-155, miR-181a, miR-451) in milk processed by temperature treatments including low temperature for long time (LTLT, 63℃ for 30 min)-, high temperature for short time (HTST, 75℃ for 15 s)-, and ultra heat treatment (UHT, 120-130℃ for 0.5-4 s). All targeted miRNAs had significantly reduced levels in processed milks compared to colostrum and raw mature milk. Interestingly, the amount of immune-related miRNAs from HTST milk was more resistant than those of LTLT and UHT milks. Our present study examined defined methods of RNA isolation and quantification of immune-specific miRNAs from small volumes of milk for use in further analysis.

Effects of Soy Isoflavones on Lipid Profiles and Hepatic LDL Receptor mRNA Level in Growing Female Rats

  • Jo, Hyun-Ju;Choi, Mi-Ja;Yoo, Min
    • Nutritional Sciences
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    • v.9 no.2
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    • pp.74-81
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    • 2006
  • The present study examined the effect of soy isoflavones on lipid metabolism in growing female rats. Rats were randomly assigned to three different groups and provided experimental diets for 9 weeks. The experimental groups were classified into 1) a control group, 2) a soy protein isolate group: soy (+)) group and 3) a soy protein concentration group: soy (-)) group. Diets contained either casein or one of two soy proteins with (soy (+)) or without isoflavones (soy (-)). Serum triglyceride concentration showed no significant differences among the experimental groups. Serum total cholesterol concentration was significantly lower in both the soy (+) and soy (-) groups than in the control group and LDL-cholesterol concentration was significantly lower in the soy (+). Serum HDL-cholesterol concentration was significantly higher in the control group than in the soy protein groups but the HDL-cholesterol share rate in total cholesterol tended to be lower in the control group than in the soy protein groups, insignificant as it was. Hepatic IDL receptor mRNA level was significantly increased in the soy (+) group when compared to the other two groups to be 20% higher than the control group. In conclusion, soy protein isolate, soy protein rich with isoflavones reduced serum total cholesterol and LDL-cholesterol concentration and increased hepatic IDL receptor mRNA expression in growing female rats. Therefore, it is considered that the intake of soy isoflvones during puberty can be advantageous in terms of the long-tenn control of serum lipid.

Studies on the Chromatin Isolated from the Organs of Animals Received Whole-body X-ray Irradiation (백서장기(白鼠臟器)에서의 Chromatin의 분리(分離)와 그 RNA 합성능(合成能)에 미치는 X-선전신조사(線全身照射)의 영향(影響)에 관(關)한 연구(硏究))

  • Han, Su-Nam
    • The Korean Journal of Nuclear Medicine
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    • v.1 no.2
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    • pp.27-34
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    • 1967
  • 1. Within experimental chromatin, the total protein: DNA ratio did not vary in the same organs of control and irradiated rats. However, the amount of RNA and total protein associated with the DNA varied considerably among the different types of chromatin. In particular, the content of chromatin was the highest in the irradiated tissue, and the lowest in the chromatin control tissue. RNA and total protein ratio of chromatins from brain, liver, testis and spleen declined with experimental organs. 2. There was the same quantitative relationship between the amount of RNA and the amount histone-protein associated with DNA in each chromatin. 3. RNA:DNA ratio of chromatin showed a $1.5{\sim}2$ times increase in the irradiated organs except brain. However, RNA:DNA ratio was decreased in chromatin by irradiation. 4. Histone-protein:Residual protein ratio was greatly varied among the organs. However, the effect was not found by irradiation. 5. Priming activity of chromatins showed a higher value in testis and the activity was greater in organs with higher metabolic activity. 6. Inhibition of Actinomycin D observable in chromatin for testis, liver, spleen and brain declined without relationship between irradiated and non-irradiated conditions. Ammonium sulfate in DNA of chromatin from histone showed increased priming activity with dissociation by Electrostatics. It may give different effect of ammonium sulfate on stimulation by property of chromatins. 7. It is suggested that the results support a proposal that the higher sensitivity of radioactive in testis, spleen by irradiated showed a increase and decrease lower-sensitivity of radioactive from brain, liver than did priming activity under the radioactive conditions.

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Effects of Isotlavones Supplemented Diet on Lipid Concentrations and Hepatic LDL Receptor mRNA Level in Growing Female Rats (성장기 암컷 흰쥐에서 이소플라본 첨가 식이가 지질 농도와 간 LDL 수용체의 유전자 발현정도에 미치는 영향)

  • Choi Mi-Ja;Jo Hyun-Ju
    • Journal of Nutrition and Health
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    • v.38 no.5
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    • pp.344-351
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    • 2005
  • The purpose of this study was to examine the impact of isoflavones on lipid concentrations and hepatic LDL receptor mRNA level in growing female rats. Twenty four rats (body weight $75\pm5g$) were randomly assigned to one of two groups, consuming control diet or isoflavones supplemented diet (57mg isoflavones/100g diet). All rats has been fed on experimental diet and deionized water ad libitum for 9 weeks. The concentration of triglyceride and total cholesterol were measured in serum and liver. Serum HDL cholesterol was measured. Hepatic LDL receptor mRNA level was tested by RT-PCR. Supplementation of isoflavones did not affect weight gain, mean food intake and food efficiency ratio. Serum total cholesterol and non-HDL cholesterol of isoflavones supplemented rats were significantly lower than those of control rats (p<0.05). But hepatic cholseterol was not influenced by supplementation of isoflavones. Hepatic LDL receptor mRNA level not significantly different between control group and isoflavones supplemented group. Therefore, isoflavones may be beneficial on serum cholesterol and non-HDL cholesterol lowering in growing female rats.

Biochemical Changes of Protein during the Senescence of Tobacco Leaf (담배잎의 노화과정에 따른 단백질의 생화학적 변화)

  • 이상각;심상인;강병화
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.41 no.5
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    • pp.563-568
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    • 1996
  • This experiment was conducted to obtain basic information of biochemical changes in the process of senescence by measuring the total RNA, protein, protease activity and electrophoretic pattern of protein in tobacco plant. The content of soluble protein increased by 15 days after leaf emergence and its level was not changed from 15 to 35 days after leaf emergence. The content of total RNA showed a maximum value at 15 days after leaf emergence and then decreased rapidly until 30 days after leaf emergence. The activity of protease of neutral fraction was higher than that of acidic fraction and rapidly increased up to the end of senescence after 50 days after leaf emergence. According to the analysis of electrophoresis, polypeptide band of 61kd was developed after 35 days after leaf emergence and increased by the end of senescence.

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Optimal RNA Extraction Methods and Development of Synthetic Clones for Seven Strawberry Viruses (딸기바이러스 진단을 위한 최적의 RNA 추출 방법 및 주요 7종 딸기바이 러스의 진단법 개발)

  • Kwon, Sun-Jung;Yoon, Ju-Yeon;Cho, In-Sook;Chung, Bong-Nam
    • Research in Plant Disease
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    • v.26 no.3
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    • pp.170-178
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    • 2020
  • Most strawberry viruses exist relatively low titers in tissues, and strawberry tissues include high levels of contamination by polysaccharides and phenolic compounds. These traits make the efficiency of strawberry diagnosis difficult. In this study, we tested different commercially available kits and reagents to secure optimal RNA extraction methods to determine virus detection from strawberry leaves. Total RNA was isolated from leaves of strawberry mottle virus (SMoV)-infected strawberry cultivar 'Mihong'. The efficiency of total RNA for virus diagnosis was confirmed through SMoV detection by one-step or two-step reverse transcription and polymerase chain reaction (RT-PCR). Among those, the RNeasy plant RNA kit was best to isolate RNA and the isolated RNA was good enough for further applications. To ensure a reliable detection for strawberry viruses, synthetic diagnosis clones for major seven strawberry viruses such as strawberry mild yellow edge virus, SMoV, strawberry latent ring spot virus, strawberry crinkle virus, strawberry pallidosis associated virus, strawberry vein banding virus and strawberry necrotic spot virus have been constructed. Based on the synthetic genes in each clone, primer sets for seven strawberry viruses were designed and tested an RT-PCR condition through a simultaneous application of the same annealing temperature that allowed to achieve an efficient and convenient diagnosis.

Glucose Transporter Gene Expression in Human Lung Cancer Cell Lines (사람 폐암 세포주에서 포도당 운반 단백 유전자의 발현)

  • Kim, Woo-Jin;Yim, Jae-Joon;Lee, Jae-Ho;Yoo, Chul-Gyu;Chung, Hee-Soon;Han, Sung-Koo;Chung, June-Key;Shim, Young-Soo;Kim, Young-Whan
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.4
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    • pp.760-765
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    • 1998
  • Background: Glucose uptake has been found to be increased in cancer cells, and FDG-PET imaging is used for diagnosis of cancer using this phenomenon. However, the exact mechanism of increased glucose uptake in cancer cells has not been clarified. Recent studies demonstrated the presence of glucose transporter(GLUT) mRNA expression in gastrointestinal cancer and head and neck cancer, and suggested that GLUT may be associated with glucose uptake in cancer cells. In lung cancer cells, glucose metabolism is also known to be increased. We evaluated GLUT mRNA expression in human lung cancer cell lines in order to find out the mechanism of increased glucose uptake in lung cancer. Method: Total RNA was isolated from 15 human lung cancer cell lines and immortalized bronchial epithelial cell line(BEAS-2B). After electrophoresis of $20{\mu}g$ total RNA, Northern blot analysis was done using GLUT1 cDNA and GLUT3 cDNA as probes. Results: Thirteen of 14 human lung cancer cell lines expressed GLUT1 mRNA and 10 of 14 human lung cancer cell lines expressed GLUT3 mRNA. Eight human lung cancer cell lines expressed both GLUT mRNAs. BEAS-2B expressed GLUT1 mRNA and did not express detectable GLUT3 mRNA. Conclusion: The increase of glucose metabolism in lung cancer may be associated with GLUT1 and GLUT3 expression.

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Acute Pulmonary Responses in Vivo to Silica Complexed with $H^+$, $Zn^{2+}$, or $Fe^{3+}$

  • Lee, Ji-Hee
    • The Korean Journal of Physiology and Pharmacology
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    • v.3 no.2
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    • pp.183-189
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    • 1999
  • This investigation is to determine whether the surface complexation of iron influence acute pulmonary responses induced by silica. For this study, three varieties of cation complexed silica were used: $silica-H^+,\;-Zn^{2+},\;and\;-Fe^{3+},$ since the first two are not active in the transport of electrons and generate little free radicals relative to the dust with the surface iron. Rats (270 to 280 g) were intratracheally (IT) instilled with saline, $silica-H^+,\;-Zn^{2+},\;or\;-Fe^{3+}$(5 mg in 0.5 ml saline). After 4 h, cell number, type, and differentiation were analysed in the bronchoalveolar lavage cells, and the levels of lactate dehydrogenase (LDH) and total protein were determined in the lavage fluid. In addition, bronchoalveolar lavage cells were cultured, and nitric oxide production was measured using nitrate assay. Inducible nitric oxide synthase (iNOS) mRNA in the bronchoalveolar lavage cells was also determined by northern blot analysis. Differential counts of the lavage cells showed that red blood cells were increased by 9-, 8-, and 13-fold and total leukocytes (lymphocytes plus polymorphonuclear neutrophils) by 48-, 36-, and 33-fold, following IT $silica-H^+,\;-Zn^{2+},\;and\;-Fe^{3+},$ respectively compared with the saline group. Meanwhile, there were no significant differences in red blood cells and total leukocytes among any of the cation complexed silica groups. The levels of LDH and total protein in the lavage fluid were significantly increased by 3- to 4-fold. However, compared among these silica groups, $Fe^{3+}$? complexation did not significantly change the LDH activity and total protein. NO production in cultured bronchoalveolar lavage cells was elevated by 2-fold, following IT any of the silica treatments compared with the saline group. Furthermore, the steady-state levels of iNOS mRNA in the lavage cells were greatly increased. There were any differences in iNOS mRNA expression among the silica-treated groups as with NO production. These findings suggest that surface complexed iron may not influence the acute pulmonary responses resulted from 4h exposure to silica.

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