• Title/Summary/Keyword: Differential Screening

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Identification of Differentially Expressed Genes in Human Small Cell Lung Carcinoma Using Subtractive Hybridization

  • Ahn Seung-Ju;Choi Jae-Kyoung;Joo Young Mi;Lee Min-A;Choi Pyung-Rak;Lee Yeong-Mi;Kim Myong-Shin;Kim So-Young;Jeon Eun-Hee;Min Byung-In;Kim Chong-Rak
    • Biomedical Science Letters
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    • v.10 no.3
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    • pp.195-202
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    • 2004
  • Lung cancer is a leading cause of cancer death worldwide; however, despite major advances in cancer treatment during the past two decades, the prognostic outcome of lung cancer patients has improved only minimally. This is largely due to the inadequacy of the traditional screening approach of diagnosis in lung cancer, which detects only well­established overt cancers and fails to identify precursor lesions in premalignant conditions of the bronchial tree. In recent years this situation has fundamentally changed with the identification of molecular abnormalities characteristic of premalignant changes; these concern tumour suppressor genes, loss of heterozygosity at crucial sites and activation of oncogenes. Basic knowledge at the molecular level has extremely important clinical implications with regard to early diagnosis, risk assessment and prevention, and therapeutic targets. In this study we used a 'cap-finder' subtractive hybridization method, 'long distance' polymerase chain reaction (PCR), streptavidin magnetic beads mediated subtraction, and spin column chromatography to detect differential expression genes of human small cell lung carcinoma. We have now isolated ninety two genes that expressed differentially in the human small cell lung carcinoma cells and analyzed of 12 clones with sequencing, nine cDNAs include tapasin (NGS-17) mRNA, BC200 alpha scRNA, chromosome 12q24 PAC RPCI3-462E2, protein phosphatase 1 (PPPICA), translocation protein 1 (TLOC1), ribosomal protein S24 (RPS24) mRNA, protein phosphatase (PPEF2), cathepsin Z, MDM2 gene and three novel genes. They may be oncogenesis­related proteins.

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Screening of Differentially Expressed Genes between PC12 Cells and A123.7 Cells (PC12 세포와 A123.7 세포에서 차별적으로 발현되는 유전자의 검색)

  • Baik, Seung-Youn;Yang, Byung-Hwan;Chai, Young-Gyu
    • Korean Journal of Biological Psychiatry
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    • v.6 no.1
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    • pp.67-73
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    • 1999
  • The cAMP-dependent protein kinase(PKA) is an intracellular enzyme with serine-threonine kinase activity that plays a key role in cell growth, differentiation, and apoptosis in eukaryotes. In order to understand the PKA signal transduction pathway regulating cell life cycle and identify its role, we focused on the characterization of up-/down-regulated genes by PKA using the differential display polymerase chain reaction. Seven differentially expressed sequence tags(DEST) have been obtained. Among these DESTs, 2 DESTs were homologous to the sequence of genes from BLAST search result. KC1-5 DEST that was up-regulated in A123.7 cells was highly corresponded to mouse apoptosis-related gene(MA-3) or mouse mRNA for topoisomerase inhibitor suppressed(TIS). MA-3 was induced in various types of apoptosis, specially in NGF-deprived apoptotic PC12 cells. TIS was down-regulated in the RVC lymphoma cells incubated with topoisomerase inhibitor that induces DNA strand breakages. PG1-1 DEST that was highly expressed in PC12 cells was corresponded to transposon Tn10 3'-end. Tnansposon Tn10 was up-regulated in differentiated myeloblastic ML-1 cells by 12-O-tetradecanoylphorbol-13-acetate. This study illuminates that MA-3/TIS was down-regulated by PKA activity, and transposon Tn10 was up-regulated by it.

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Genotypic Identification of Cystoisospora in Immunocompromised Patients Using Tm-Variation Analysis

  • Basyoni, Maha M.A.;Elghobary, Hany Ahmed Fouad
    • Parasites, Hosts and Diseases
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    • v.55 no.6
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    • pp.601-606
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    • 2017
  • Cystoisospora is responsible for morbidity in immunocompromised patients. PCR is sensitive for diagnosing Cystoisospora; however, it needs reevaluation for differential molecular diagnosis of cystoisosporiasis. We aimed at evaluating melting curve analysis (MCA) after real-time PCR (qPCR) in diagnosis and genotyping of Cystoisospora as an alternative to conventional PCR. We included 293 diarrheic stool samples of patients attending the Department of Clinical Oncology and Nuclear Medicine of Cairo University Hospitals, Egypt. Samples were subjected to microscopy, nested PCR (nPCR), and qPCR targeting the internal transcribed spacer 2 region (ITS2) of the ribosomal RNA (r RNA) gene followed by melting temperatures ($T_ms$) analysis and comparing the results to PCR-RFLP banding patterns. Using microscopy and ITS2-nPCR, 3.1% and 5.8% of cases were Cystoisospora positive, respectively, while 10.9% were positive using qPCR. Genotyping of Cystoisospora by qPCR-MCA revealed 2 genotypes. These genotypes matched with 2 distinct melting peaks with specified $T_ms$ at $85.8^{\circ}C$ and $88.6^{\circ}C$, which indicated genetic variation among Cystoisospora isolates in Egypt. Genotype II proved to be more prevalent (65.6%). HIV-related Kaposi sarcoma and leukemic patients harbored both genotypes with a tendency to genotype II. Genotype I was more prevalent in lymphomas and mammary gland tumors while colorectal and hepatocellular tumors harbored genotype II suggesting that this genotype might be responsible for the development of cystoisosporiasis in immunocompromised patients. Direct reliable identification and differentiation of Cystoisospora species could be established using $qPCR-T_ms$ analysis which is useful for rapid detection and screening of Cystoisospora genotypes principally in high risk groups.

Resistance of Plants to Herbicide (제초제(除草劑)에 대한 식물(植物)의 저항성(抵抗性))

  • Kim, Kil-Ung
    • Korean Journal of Weed Science
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    • v.4 no.1
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    • pp.96-106
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    • 1984
  • Changes in weed floras and development of plant resistance to herbicides seemed to be closely related with increased and repeated use of herbicides. Herbicide use increased from 5% of the total consumption of pesticide in 1950 to 45% in 1976 in world basis. About 200 herbicides have been introduced to agriculture so as to control about 206 weed species which have been recorded important to human beings. In Korea, there was about 351 times in increased use of herbicides from 1966 to 1982. Interspecific selection by herbicide is mainly responsible for changes in weed floras and resulted in varying tolerance or susceptibility to herbicides, together with the changes of agricultural practices. The present trend toward continuous cereal cultivation throughout world will lead to type of changes in weed floras favorable to therophyte which can survive under unfavorable conditions as seeds rather than the types of geophyte which can survive unfavorable seasons as buds placed below soil surface. However, geophyte such as Sagitaria pygmaea, and Scirpus jurtcoides, and Cyperus rotundus and Cynodon dactylon in temperate warm climate become severe paddy weeds, presumably because of the removal of annual weeds by herbicides. Since differential tolerance to 2,4-D was firstly reported in Agrostis stolofera, about 30 species of weeds in 18 genera are presently known to have developed resistance to triazine herbicides. Resistance of weed biotypes to triazine herbicide is not mainly due to limited absorption and translocation or to the difference in metabolism, but is the result of biochemical changes at the site of metabolic activity, such as a loss of herbicide affinity for triazine binding site in the photosystem II complex of the chloroplast membrane. Genetical study showed that plastid resistance to triazine was wholly inherited through cytoplasmic DNA in the case of Brassica campestris. Plant tissue culture method can be utilized as an alternate mean of herbicide screening and development of resistance variants to herbicides as suggested by Chaleff and Parsons. In this purpose, one should be certain that the primary target process is operational in cell culture. Further, there are a variety of obstacles in doing this type of research, particularly development of resistance source and it's regeneration because cultured cells and whole plants represent different developmental state.

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Molecular cloning of peroxidase cDNAs from dehydration-treated fibrous roots of sweetpotato and their differential expression in response to stress

  • Kim, Yun-Hee;Yang, Kyoung-Sil;Kim, Cha-Young;Ryu, Sun-Hwa;Song, Wan-Keun;Kwon, Suk-Yoon;Lee, Haeng-Soon;Bang, Jae-Wook;Kwak, Sang-Soo
    • BMB Reports
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    • v.41 no.3
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    • pp.259-265
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    • 2008
  • Three peroxidase (POD) cDNAs were isolated from dehydration-treated fibrous roots of sweetpotato (Ipomoea batatas) plant via the screening of a cDNA library, and their expressions were assessed to characterize functions of each POD in relation to environmental stress. Three PODs were divided into two groups, designated the basic PODs (swpb4, swpb5) and the anionic PODs (swpa7), on the basis of the pI values of mature proteins. Fluorescence microscope analysis indicated that three PODs are secreted into the extracellular space. RT-PCR analysis revealed that POD genes have diverse expression patterns in a variety of plant tissues. Swpb4 was abundantly expressed in stem tissues, whereas the expression levels of swpb5 and swpa7 transcripts were high in fibrous and thick pigmented roots. Swpb4 and swpa7 showed abundant expression levels in suspension cultured cells. Three POD genes responded differently in the leaf and fibrous roots in response to a variety of stresses including dehydration, temperature stress, stress-associated chemicals, and pathogenic bacteria.

Non-neoplastic Myelopathies Mimicking Intramedullary Spinal Cord Tumors : Retrospective Analysis of 8 Surgically Proven Cases (척수내 종양과 감별을 요하는 비종양성 척수증 : 수술로 확진된 8례의 후향적 분석)

  • Kim, Ki-Jeong;Chung, Chun-Kee;Sim, Ki-Bum;Kim, Hyun-Jib
    • Journal of Korean Neurosurgical Society
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    • v.29 no.7
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    • pp.891-898
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    • 2000
  • Objective : It is difficult to differentiate intramedullary spinal cord tumors preoperatively from non-neoplastic pathologies in patients presenting as non-compressive myelopathies in magnetic resonance imaging(MRI). In this report, the authors reviewed nonneoplastic intramedullary spinal cord lesions preoperatively diagnosed as tumors and discussed their clinical and radiological characteristics and usefulness of surgical intervention. Methods : From January, 1985 to January, 1999, authors experienced eight non-neoplastic pathologies mimicking intramedullary spinal cord tumors and analysed their medical records, radiological findings and histopathological specimens retrospectively. Results : There were five males and three females and the duration of symptoms were from two to 20 months(mean, 9.8 months). The location of lesions were four cervical, one cervicothoracic and three thoracic. All patients manifested sensory abnormality, seven motor weakness, and six bladder symptom. All cases had swollen spinal cords and increased signal intensities in spin-echo sequences. Six cases showed contrast enhancement : four cases were focal and two diffuse. Under the impression of intramedullary tumors, the patients were operated upon. Final diagnoses on the base of clinical and pathologic finding were : three subacute necrotizing myelopathies, two multiple scleroses, two myelopathy of unknown etiology. One case showed no gross abnormality in surgical field in spite of adequate exposure of the lesion, so biopsy was not performed. In that case, postoperative MRI revealed spontaneous resolution of the lesion. Conclusion : MRI is invaluable diagnostic tool in screening myelopathies. However, its high sensitivity and lack of specificity make difficulty in preoperative differential diagnosis of non-compressive myelopathies. Although no surgical morbidity occurred in our series, we sometimes failed to confirm definite diagnosis even with biopsy. In such a circumstance, long-term follow up is needed.

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Radioisotope Perfusion Lung Scanning in Takayasu's Arteritis (Takayasu 동맥염에서 폐관류스캔에 관한 연구)

  • Park, Seok-Gun;Bom, Hee-Seung;Oh, Yeon-Sang;Han, Jin-Suk;Chung, June-Key;Choi, Sung-Jae;Kim, Byung-Kook;Lee, Jung-Sang;Lee, Young-Woo;Koh, Chang-Soon;Lee, Mun-Ho
    • The Korean Journal of Nuclear Medicine
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    • v.20 no.2
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    • pp.73-78
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    • 1986
  • Of 18 patients with Takayasu's arteritis who underwent $^{99m}Tc-MAA$ (macroaggregated albumin) perfusion luing scanning, 9 (50%) showed perfusion defects. Chest X-rays of them were nonspecific. One patient underwent $^{99m}Tc-DTPA$ radio aerosol inhalation lung scanning simultaneously, which revealed normal. So routine screening radioisotope perfusion lung scanning is helpful to evaluate pulmonary arterial involnmement in Takayasu's arteritis patients. And Takayasu's arteritis should be included in differential diagnosis of pulmonary embolism.

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Differential Proteomic Analysis of Secreted Proteins from Cutinase-producing Bacillus sp. SB-007

  • Ban, Yeon-Hee;Jeon, Mi-Ri;Yoon, Ji-Hee;Park, Jae-Min;Um, Hyun-Ju;Kim, Dae-Soon;Jung, Seung-Ki;Kim, Keun-Young;Lee, Jee-Won;Min, Ji-Ho;Kim, Yang-Hoon
    • The Plant Pathology Journal
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    • v.24 no.2
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    • pp.191-201
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    • 2008
  • Bacillus sp. SB-007 was isolated from pea leaves harvested from the southwestern parts of South Korea through screening on a minimal medium containing 0.2% purified cutin for its ability to induce the cutinase production. However, no cutinase was produced when it was grown in a minimal medium containing 0.2% glucose. A proteomic approach was applied to separate and characterize these differentially secreted proteins. The expression level of 83 extracellular proteins of the cutinase-producing Bacillus sp. strain SB-007 incubated in a cutinase-induced medium increased significantly as compared with that cultured in a non cutinase-induced medium containing glucose. The extracellular proteome of Bacillus sp. SB-007 includes proteins from different functional classes, such as enzymes for the degradation of various macromolecules, proteins involved in energy metabolism, sporulation, transport/binding proteins and lipoproteins, stress inducible proteins, several cellular molecule biosynthetic pathways and catabolism, and some proteins with an as yet unknown function. In addition, the two protein spots showed little similarities with the known lipolytic enzymes in the database. These secreted proteome analysis results are expected to be useful in improving the Bacillus strains for the production of industrial cutinases.

Screening of toxic potential of graphene family nanomaterials using in vitro and alternative in vivo toxicity testing systems

  • Chatterjee, Nivedita;Yang, Ji Su;Park, Kwangsik;Oh, Seung Min;Park, Jeonggue;Choi, Jinhee
    • Environmental Analysis Health and Toxicology
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    • v.30
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    • pp.7.1-7.7
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    • 2015
  • Objectives The widely promising applications of graphene nanomaterials raise considerable concerns regarding their environmental and human health risk assessment. The aim of the current study was to evaluate the toxicity profiling of graphene family nanano-materials (GFNs) in alternative in vitro and in vivo toxicity testing models. Methods The GFNs used in this study are graphene nanoplatelets ([GNPs]-pristine, carboxylate [COOH] and amide [$NH_2$]) and graphene oxides (single layer [SLGO] and few layers [FLGO]). The human bronchial epithelial cells (Beas2B cells) as in vitro system and the nematode Caenorhabditis elegans as in vivo system were used to profile the toxicity response of GFNs. Cytotoxicity assays, colony formation assay for cellular toxicity and reproduction potentiality in C. elegans were used as end points to evaluate the GFNs' toxicity. Results In general, GNPs exhibited higher toxicity than GOs in Beas2B cells, and among the GNPs the order of toxicity was pristine > $NH_2$ > COOH. Although the order of toxicity of the GNPs was maintained in C. elegans reproductive toxicity, but GOs were found to be more toxic in the worms than GNPs. In both systems, SLGO exhibited profoundly greater dose dependency than FLGO. The possible reason of their differential toxicity lay in their distinctive physicochemical characteristics and agglomeration behavior in the exposure media. Conclusions The present study revealed that the toxicity of GFNs is dependent on the graphene nanomaterial's physical forms, surface functionalizations, number of layers, dose, time of exposure and obviously, on the alternative model systems used for toxicity assessment.

The Effect of Dietary Docosahexaenoic Acid Enrichment on the Expression of Porcine Hepatic Genes

  • Chang, W.C.;Chen, C.H.;Cheng, W.T.K.;Ding, S.T.
    • Asian-Australasian Journal of Animal Sciences
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    • v.20 no.5
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    • pp.768-774
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    • 2007
  • To study the effect of dietary docosahexaenoic acid (DHA) enrichment on the expression of hepatic genes in pigs, weaned, crossbred pigs (30 d old) were fed diets supplemented with either 2% tallow or DHA oil for 18 d. Hepatic mRNA was extracted. Suppression subtractive hybridization was used to explore the hepatic genes that were specifically regulated by dietary DHA enrichment. After subtraction, we observed 288 cDNA fragments differentially expressed in livers from pigs fed either 2% DHA oil or 2% tallow for 18 d. After differential screening, 7 genes were found to be differentially expressed. Serum amyloid A protein 2 (SAA2) was further investigated because of its role in lipid metabolism. Northern analysis indicated that hepatic SAA2 was upregulated by dietary DHA enrichment (p<0.05). In a second experiment, feeding 10% DHA oil for 2d significantly increased the expression of SAA2 (compared to the 10% tallow group; p<0.05). The porcine SAA2 full length cDNA sequence was cloned and the sequence was compared to the human and mouse sequences. The homology of the SAA2 amino acid sequence between pig and human was 73% and between pig and mouse was 62%. There was a considerable difference in SAA2 sequences among these species. Of particular note was a deletion of 8 amino acids, in the pig compared to the human. This fragment is a specific characteristic for the SAA subtype that involved in acute inflammation reaction. Similar to human and mouse, porcine SAA2 was highly expressed in the liver of pigs. It was not detectable in the skeletal muscle, heart muscle, spleen, kidney, lung, and adipose tissue. These data suggest that SAA2 may be involved in mediation of the function of dietary DHA in the liver of the pig, however, the mechanism is not yet clear.