• Title/Summary/Keyword: Cell fusion technique

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Importance of FISH combined with Morphology, Immunophenotype and Cytogenetic Analysis of Childhood/Adult Acute Lymphoblastic Leukemia in Omani Patients

  • Goud, Tadakal Mallana;Al Salmani, Kamla Khalfan;Al Harasi, Salma Mohammed;Al Musalhi, Muhanna;Wasifuddin, Shah Mohammed;Rajab, Anna
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.16
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    • pp.7343-7350
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    • 2015
  • Genetic changes associated with acute lymphoblastic leukemia (ALL) provide very important diagnostic and prognostic information with a direct impact on patient management. Detection of chromosome abnormalities by conventional cytogenetics combined with fluorescence in situ hybridization (FISH) play a very significant role in assessing risk stratification. Identification of specific chromosome abnormalities has led to the recognition of genetic subgroups based on reciprocal translocations, deletions and modal number in B or T-cell ALL. In the last twelve years 102 newly diagnosed childhood/adult ALL bone marrow samples were analysed for chromosomal abnormalities with conventional G-banding, and FISH (selected cases) using specific probes in our hospital. G-banded karyotype analysis found clonal numerical and/or structural chromosomal aberrations in 74.2% of cases. Patients with pseudodiploidy represented the most frequent group (38.7%) followed by high hyperdiploidy group (12.9%), low hyperdiploidy group (9.7%), hypodiploidy (<46) group (9.7%) and high hypertriploidy group (3.2%). The highest observed numerical chromosomal alteration was high hyperdiploidy (12.9%) with abnormal karyotypes while abnormal 12p (7.5%) was the highest observed structural abnormality followed by t(12;21)(p13.3;q22) resulting in ETV6/RUNX1 fusion (5.4%) and t(9;22)(q34.1;q11.2) resulting in BCR/ABL1 fusion (4.3%). Interestingly, we identified 16 cases with rare and complex structural aberrations. Application of the FISH technique produced major improvements in the sensitivity and accuracy of cytogenetic analysis with ALL patients. In conclusion it confirmed heterogeneity of ALL by identifying various recurrent chromosomal aberrations along with non-specific rearrangements and their association with specific immunophenotypes. This study pool is representative of paediatric/adult ALL patients in Oman.

Studies on nuclear transplantation in mouse embryos. I. Functional differences between maternal and paternal genomes (생쥐 수정란의 핵이식에 관한 연구 I. 모성 및 부성 genome의 기능차이에 관한 연구)

  • Choe, Sang-yong;Park, Choon-saeng;Lee, Hyo-jong;Park, Hee-sung
    • Korean Journal of Veterinary Research
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    • v.30 no.2
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    • pp.123-127
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    • 1990
  • By nuclear transplantation technology twenty eight mice have been produced after transfer of heterozygous biparental eggs. Also heterozygous gynogenetic eggs with two female pronuclei and heterozygous androgenetic eggs with two male pronuclei have been obtained by injecting a male or female pronucleus with Sendai virus into the perivitelline space of enucleated haploid zygotes at pronuclear stage. The success rate of enucleation, karyoplast injection and fusion of both the pronuclei was 80.3, 83.4 and 81.8%, respectively. The overall pronuclei fusion rates by this technique were 56, 50 and 56% in biparental, gynogenetic and androgenetic eggs, respectively. The evidence was ascertained that the gynogenetic and androgenetic eggs were also able to develop in vitro up to blastocyst stage, even though their developmental potential was greatly diminished beyond 2-cell stage. The gynogenetic eggs were able to develop in vivo up to day 10 of pregnancy, while the androgenetic eggs failed to develop in vivo during the same period.

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Study on the Improvement of Bovine Somatic Cell Nuclear Transfer Technique (소 체세포 핵이식기술의 효율 증진에 관한 연구)

  • 양윤희;최종엽;이상영;박춘근;양부근;김정익;정희태
    • Korean Journal of Animal Reproduction
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    • v.27 no.3
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    • pp.233-240
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    • 2003
  • This study was conducted to examine the effects of oocyte maturation period, phytohemagglutinin-P (PHA-P) treatment and activation agent on the enucleation, fusion, activation or in vitro development of bovine nuclear transfer embryos. Bovine oocytes were enucleated at 16∼24 h of in vitro maturation (IVM). Adult ear skin cells treated or non-treated with PHA-P were transferred into enucleated oocytes. Reconstituted oocytes treated or non-treated with PHA-P were fused by a pulse of 1.5 kV/cm for 30 $\mu$sec. Fused oocytes were activated with a combination of calcium ionophore (A23187) and cycloheximide (CHXM) or dimethylaminopurine (DMAP), and cultured in vitro for 7∼9 days. Enucleation rate was significantly increased when oocytes were matured for 16∼18 h (70.2∼92.3%, P<0.05) compared to that of oocytes were matured for 20∼24 h (44.3∼53.4%). The location of metaphase-II plate was far off from the 1st polar body as maturation time was increased. PHA-P treatment of donor cells or reconstituted oocytes significantly improved fusion rate (P<0.05). Cleavage and blastocyst formation rates were significantly increased after activation with a combination of A23187 and DMAP (78.6% and 32.9%, respectively) compared to those of embryos activated with a combination of A23l87 and CHXM (48.5 and 15.2%, respectively). From the present result, it is suggested that high enucleation efficiency can obtained by using oocytes matured for 18 h. It also shows that PHA-P treatment can improve the fusion rate, and activation with a combination of A23187 and DMAP can enhance the embryo development.

Study of Trend of Reporting on GMO as a Convergence Technology: Focused on analyzing Articles in Major Domestic Daily Newspapers from 1994 to 2015 (융복합기술로서 GMO에 관한 보도경향연구: 1994~2015년까지 국내 주요일간지 기사분석을 중심으로)

  • Cho, Hang-Min
    • Journal of Digital Convergence
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    • v.14 no.12
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    • pp.267-281
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    • 2016
  • This study analyzed the contents of articles on GMO in Chosun Ilbo and Hankyoreh from 1994 through 2015 to check the trend of reporting on GMO as a science and technology risk issue. As a result, 'risk' and 'anxiety/concern' were continuously treated as important subjects, and there was a lack of depth and journalist's professionalism in all three periods: 'Introduction of GMO technology (1994-2000)', 'Development of GMO technology (2001-2010)' and 'Social acceptance of GMO (2011-2015).' In a comparison between the media with different ideological goals, the Chosun Ilbo had a stronger new-technological inclination to GMO than the Hankyoreh in reporting topics and tones of arguments. As a result of checking the relation of the tones in reporting with the 'risk-benefit' of GMO, reports in negative tones underlined 'risk' while those in positive tones, 'benefit.' This study could suggest improvements of subject bias and unprofessionalism in the domestic science journalism, the barometer of public awareness of disputes over science and technology risk.

Recolonization of Transfected Blastodermal Cells in Developing Embryos after Transferring into UV-irradiated Fertilized Hen′s Egg (UV-조사 수정란 내로 이식한 유전자 변화 배반엽 세포의 재구성)

  • Lee, K.S.;Lee, H.;Kim, K.D.;Park, S.S.;Lee, S.H.
    • Korean Journal of Poultry Science
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    • v.27 no.2
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    • pp.155-161
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    • 2000
  • Unfortunately, there is no technique which is stable and repetitive to produce transgenic chicken, although various ways of gene transfer including PGC-and embryonic cell-mediated gene transfer, DNA microinjection, virus inoculation and sperm cells have been employed. The aims of this study were 세 develop and establish such a stable, repetitive and efficient way of gene transfer giving a faithful gene expression during development after the reconstruction of embryo in an UV-irradiated egg. A dual reporter plasmid (pJJ9), a fusion gene containing lacZ and GFP driven by a CMV promoter was used to exploit either merits of both reporting markers. lacZ with strong signal or GFP with vital marking. Electroporated embryonic blastodermal cells (EBCs) in the presence of the pJJ9 DNA faithfully showed 377 bp PCR product and lacZ or GFP expressions in the identical cells in vitro of in vivo. Furthermore, analyses of expression pattern of the foreign DNA demonstrated that microinjected EBCs cells into the UV-irradiated recipient egg should participate in normal developmental process, for example, proliferation and differentiation into various tissues. Thirty percentages of the manipulated eggs showed lacZ expression in their tissues. These results together with the specific procedures used in this study should facilitate avian transgenesis.

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Analysis of the microstructure of melting-pool in aluminum specimens fabricated by SLM technique (SLM 기법으로 제작한 알루미늄 시편 내부 멜팅풀 미세조직 분석)

  • Kim, Moo-Sun
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.21 no.12
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    • pp.115-119
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    • 2020
  • Selective Laser Melting (SLM) technology is state-of-the-art additive manufacturing process technology that produces a three-dimensional structure by irradiating a laser on a fine metal powder to perform the fusion of a specific area and repeat this process. Owing to the characteristics of the additive manufacturing process, the melting phenomenon of the metal material by the laser has directionality depending on the process conditions, such as the irradiation direction of the laser and the build-up direction. For this reason, the composition of the metal material in the structure exhibits non-uniform characteristics. In this study, aluminum (AlSi10Mg) specimens were manufactured by applying SLM technology, and the material composition characteristics of the specimen were analyzed. The specimens were manufactured as cylinders by the build-up orientation of 0°, 45°, and 90°. The surface morphology of the specimen plane was analyzed optically. TEM analysis was performed on the core and the interface of the melting-pool inside the specimen generated by laser irradiation. The analysis results confirmed that there was a difference between the nano cell structure of the core and the interface of the melting-pool, and that the composition ratio of Si appeared higher at the interface than at the core of the cell.

Development of Human Antibody Inhibiting RNase H Activity of Polymerase of Hepatitis B Virus Using Phage Display Technique (Phage Display 기법을 이용한 B형 간염 바이러스 Polymerase의 RNase H 활성을 억제하는 인간 단세포군 항체의 개발)

  • Lee, Seong-Rak;Song, Eun-Kyoung;Jeong, Young-Joo;Lee Young-Yi;Kim, Ik-Jung;Choi, In-Hak;Park, Sae-Gwang
    • IMMUNE NETWORK
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    • v.4 no.1
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    • pp.16-22
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    • 2004
  • Background: To develop a novel treatment strategy for hepatitis B virus infection, a major cause of liver chirosis and cancer, we aimed to make human monoclonal antibodies inhibiting RNase H activity of P protein playing in important role in HBV replication. In this regard, phage display technology was employed and demonstrated as an efficient cloning method for human monoclonal antibody. So this study analysed the usability of human monoclonal antibody as protein based gene therapy. Methods: RNase H of HBV was expressed as fusion protein with maltose binding protein and purified with amylose resin column. Single chain Fv (scFv) phage antibody library was constructed by PCR cloning using total RNAs of PBMC from 50 healthy volunteers. Binders to RNase H were selected with BIAcore 2000 from the constructed library, and purified as soluble antibody fragment. The affinity and sequences of selected antibody fragments were analyzed with BIAcore and ABI automatic sequencer, respectively. And finally RNase H activity inhibiting assay was carried out. Results: Recombinant RNase H expressed in E. coli exhibited an proper enzyme activity. Naive library of $4.46{\times}10^9cfu$ was screened by BIAcore 2000. Two clones, RN41 and RN56, showed affinity of $4.5{\times}10^{-7}M$ and $1.9{\times}10^{-7}M$, respectively. But RNase H inhibiting activity of RN41 was higher than that of RN56. Conclusion: We cloned human monoclonal antibodies inhibiting RNase H activity of P protein of HBV. These antibodies can be expected to be a good candidate for protein-based antiviral therapy by preventing a replication of HBV if they can be expressed intracellularly in HBV-infected hepatocytes.

Analysis of Clonorchis sinensis antigens and diagnosis of clonorchiasis using monoclonal antibodies (단세포군 항체를 이용한 간흡충 항원의 분석 및 간흡충증의 진단)

  • Yong, Tae-Sun;Im, Gyeong-Il;Jeong, Pyeong-Rim
    • Parasites, Hosts and Diseases
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    • v.29 no.3
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    • pp.293-310
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    • 1991
  • Clonorchis sinensis is a common parasite of man in Korea. Researches on the specific antigens of C. sinensis would be valuable not only because those elucidate the molecular characteristics of this fluke but also because it is applicable to immunodiagnosis. Although many monoclonal antibodies have been used in the field of parasite immunology, few articles on monoclonal antibodies against C. sinensis have been published so far. The aim of this study was to analyse C. sinensis antigens recognized by monoclonal antibodies, and to set up ELISA-inhibition test using C. sinensis specific monoclonal antibodies for improved specificity of immunodiagnostic tests. By fusion between spleen cells of the mice immunized with C. sinensis water-soluble crude adult worm antigens and plasmacytoma cells of mouse origin, 29 hybridoma clones secreting anti-C. sinensis monoclonal antibodies were made, and 8 clones among those were found specific. After cell cloning, isotypes of 6 selected specific monoclonal anti- bodies were determined to be IgGl, IgG2b and IgA. Four exposed antigenic determinants of natural infection were recognized by different specific monoclonal antibodies. By enzyme-immunoelectrotransfer blot, 10 KD, 34 KD antigenic determinants were found to be reacted with CsHyb 0714-20, CsHyb 0605-10 monoclonal antibodies, respectively, The antigenic determinant recognized by CsHyb 0714-20 monoclonal antibody was revealed to be located at the surface and parenchyme of a parasite by indirect immunoauorescent antibody technique, and those reacted with CsHyb 0605-10, CsHyb 0714-25 monoclonal antibodies were found at the parenchyme and intestine. The antigenic determinant reacted with CsHyb 0605-23 monoclonal antibody was found mainly around the uterine eggs. Four antigenic determinants recognized by specific monoclonal antibodies were all found to be present in the early eluted fractions of C. sinensis antigens separated by Sephadex G-200 gel filtration. By conventional ELISA, 75% of clonorchiasis cases were found positive, but 7.1% of normal controls and 37.5% of paragonimiasis cases showed false positives. However, by ELISA-inhibition test using C. sinensis specific monoclonal antibody (CsHyb 0605-23), 77.1% of clonorchiasis cases were found positive, and there were no false positives in normal controls or paragonimiasis cases, indicating 100% specificity. The ELISA- inhibition test using monoclonal antibodies was found to have same sensitivity and definitely high specificity in comparison with conventional ELISA for serodiagnosis of human clonorchiasis.

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