• Title/Summary/Keyword: Cell sorting

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Enhanced Production of Bacterial Cellulose in Komagataeibacter xylinus Via Tuning of Biosynthesis Genes with Synthetic RBS

  • Hur, Dong Hoon;Choi, Woo Sung;Kim, Tae Yong;Lee, Sang Yup;Park, Jin Hwan;Jeong, Ki Jun
    • Journal of Microbiology and Biotechnology
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    • v.30 no.9
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    • pp.1430-1435
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    • 2020
  • Bacterial cellulose (BC) has outstanding physical and chemical properties, including high crystallinity, moisture retention, and tensile strength. Currently, the major producer of BC is Komagataeibacter xylinus. However, due to limited tools of expression, this host is difficult to engineer metabolically to improve BC productivity. In this study, a regulated expression system for K. xylinus with synthetic ribosome binding site (RBS) was developed and used to engineer a BC biosynthesis pathway. A synthetic RBS library was constructed using green fluorescent protein (GFP) as a reporter, and three synthetic RBSs (R4, R15, and R6) with different strengths were successfully isolated by fluorescence-activated cell sorting (FACS). Using synthetic RBS, we optimized the expression of three homologous genes responsible for BC production, pgm, galU, and ndp, and thereby greatly increased it under both static and shaking culture conditions. The final titer of BC under static and shaking conditions was 5.28 and 3.67 g/l, respectively. Our findings demonstrate that reinforced metabolic flux towards BC through quantitative gene expression represents a practical strategy for the improvement of BC productivity.

Identification of a Technique Optimized for the Isolation of Spermatogonial Stem Cells from Mouse Testes

  • Han, Na Rae;Park, Hye Jin;Lee, Hyun;Yun, Jung Im;Choi, Kimyung;Lee, Eunsong;Lee, Seung Tae
    • Journal of Embryo Transfer
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    • v.33 no.4
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    • pp.327-336
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    • 2018
  • To date, there are no protocols optimized to the effective separation of spermatogonial stem cells (SSCs) from testicular cells derived from mouse testes, thus hindering studies based on mouse SSCs. In this study, we aimed to determine the most efficient purification method for the isolation of SSCs from mouse testes among previously described techniques. Isolation of SSCs from testicular cells derived from mouse testes was conducted using four different techniques: differential plating (DP), magnetic-activated cell sorting (MACS) post-DP, MACS, and positive and negative selection double MACS. DP was performed for 1, 2, 4, 8, or 16 h, and MACS was performed using EpCAM ($MACS^{EpCAM}$), Thy1 ($MACS^{Thy1}$), or GFR ${\alpha}1$ ($MACS^{GFR{\alpha}1}$) antibodies. The purification efficiency of each method was analyzed by measuring the percentage of cells that stained positively for alkaline phosphatase. DP for 8 h, $MACS^{Thy1}$ post-DP for 8 h, $MACS^{GFR{\alpha}1}$, positive selection double $MACS^{GFR{\alpha}1/EpCAM}$, and negative selection double $MACS^{GFR{\alpha}1/{\alpha}-SMA}$ were identified as the optimal protocols for isolation of SSCs from mouse testicular cells. Comparison of the purification efficiencies of the optimized isolation protocols showed that, numerically, the highest purification efficiency was obtained using $MACS^{GFR{\alpha}1}$. Overall, our results indicate that $MACS^{GFR{\alpha}1}$ is an appropriate purification technique for the isolation of SSCs from mouse testicular cells.

Characterization of Human Epidermal Stem Cells and Living Skin Equivalents

  • Kim, Dong-Seok;Youn, Sang-Woong;Choi, Hye-Ryung;Cho, Hyun-Ju;Jeon, Sang-Eun;Park, Kyoung-Chan
    • Proceedings of the PSK Conference
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    • 2002.10a
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    • pp.273.2-274
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    • 2002
  • Human epidermal keratinocytes consist of stem cells. transit amplifying cells. and postmitotic differentiating cells. Among them, stem cells playa critical role in cell renewal. wound healing. and neoplasia. However. till now, specific markers of human epidermal keratinocytes are not clearly defined. In the present study. we separated putative stem cells from other cells using fluorescence activated cell sorting (FACS). based on differences in a6-integrin and CD71 expression. (omitted)

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ESCRT, autophagy, and frontotemporal dementia

  • Lee, Jin-A;Gao, Fen-Biao
    • BMB Reports
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    • v.41 no.12
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    • pp.827-832
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    • 2008
  • Many age-dependent neurodegenerative diseases are associated with the accumulation of abnormally folded proteins within neurons. One of the major proteolytic pathways in the cell is the autophagy pathway, which targets cytoplasmic contents and organelles to the lysosomes for bulk degradation under various physiological and stressful conditions. Although the importance of autophagy in cellular physiology is well appreciated, its precise roles in neurodegeneration remain largely unclear. Recent studies indicate that components of the endosomal sorting complex required for transport (ESCRT) are important in the autophagy pathway. Reduced activity of some ESCRT subunits leads to the accumulation of autophagosomes and failure to clear intracellular protein aggregates. Interestingly, rare mutations in CHMP2B, an ESCRT-III subunit, are associated with frontotemporal dementia linked to chromosome 3 (FTD3). Mutant CHMP2B proteins seem to disrupt the fusion of autophagosomes and lysosomes in cell culture models. These findings suggest a potential mechanism for the pathogenesis of FTD3 and possibly other neurodegenerative diseases as well.

A Study on New Cell Switch Fabric for Increasing the Performance of ATM Switching Systems (ATM 교환 시스템의 성능향상을 위한 새로운 셀 스위치 구조 연구)

  • 정진태;이옥재;전병실
    • Journal of the Korean Institute of Telematics and Electronics S
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    • v.34S no.3
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    • pp.12-23
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    • 1997
  • In this paper, we propose a new cell switch fabric for increasing the performance of ATM switching systems. Proposed switching network consists of a sorting network and a routing network. Both of these are multistage networks where each stage performs a fixed permutation on the incoming lines, and then routes them through a clumn of 2x2 switching elements. It is designed for distributing inputs and parallel processing to reduce the hardware complexity and obtain high performance of switching network. The structure and the operation of th eswitching network aredescribed and the performanceof the switching network is anlyzed under uniform traffic models. In this result, though the size of proposed network is increased the large scale, it has always the same throughput as the that of genral output queueing system with N=2. So, it is found that our proposed network is appropriate for the high apeed and lrger size of ATM switching systems.

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Study on precision improvement in weight measurement of an egg for the automatic egg sorting system (계란(鷄卵) 자동(自動) 선별기(選別機)의 난중측정(卵重測定) 정밀도(精密度) 향상(向上)에 관(關)한 기초연구(基礎硏究))

  • Kim, K.D.;Bok, J.S.
    • Korean Journal of Agricultural Science
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    • v.27 no.1
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    • pp.48-53
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    • 2000
  • The various errors existing in a weight measurement system in most automatic egg sorting system available in Korean poultry farms have caused a large amount of economic losses to the egg producers. The object of this study was an importance of egg measurement system by changing both the number and the arrangements of load cells to reduce measuring errors. The results obtained were summarized as follow : 1. Four arrangements of load cells were selected as follows : layout I : Conventional one load cell method layout II : One load cell located as egg moving direction layout III : Two load cells located facing each other layout IV : Two load cells located as parallel with egg moving direction 2. The results of egg weight measurement according to four arrangements (Layout I, II, III, IV) showed that the average errors were 1.1218g, 0.5953g, 0.7786g, 0.2793g respectively. This indicated that the Layout IV (measuring by 2 load cells located parallel with the egg moving direction) caused the lowest average error and the best in precision. 3. The average vibration of axis X, y, Z were resulted as $5.1937{\times}10^{-3}G$, $9.3604{\times}10^{-3}G$, and $16.8657{\times}10^{-3}G$ respectively when sorting large sized egg. This indicated that the vibration of axis-Z was relatively higher than those of axis-X, and axis-Y.

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The Electrode Shape for the Efficient Separation of Cell in Dielectrophoresis-Activated Cell Sorter (유전영동을 이용한 입자분리기의 효율적인 분리를 위한 전극 형태)

  • An, Jae-Min;Chae, Seung-Yeub;Park, Seok-Ho;Kim, Byung-Kyu
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.33 no.1
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    • pp.49-55
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    • 2009
  • This paper presents the optimal shape of microelectrode that generates dielectrophoretic(DEP) force to separate particles in homogeneous medium. The principle of the particles sorting is based on the use of the relative strengths of negative DEP (nDEP) and drag forces, as in a general DEP-activated cell sorter (DACS). To numerically calculate the DEP force and drag force, the simulation is implemented in MATLAB 7.0. The properties of particles, which are used in simulation, are similarly selected as those of cells to apply cell separation. The most optimized shape of electrode is selected by numerical simulation according to a variety of electrode shape such as rectangle, trapezoidal, and right-triangle. Through, in addition, parameter study, we found that applied frequency is more significant factor on the separation than various parameters, such as applied voltage and permittivity of medium, that decide on the strength of DEP force.

A Study for reduction of the power loss of PV modules (PV moudule의 출력손실 저감요인 분석)

  • Lee, Sang-Hun;Kang, Gi-Hwan;Yu, Gwon-Jong;Ahn, Hyung-Keun;Han, Deuk-Young
    • 한국태양에너지학회:학술대회논문집
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    • 2011.11a
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    • pp.45-50
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    • 2011
  • The efficiency of solar cell was about 4[%] in initial stage of photovoltaic industry, but it has quite a lot of efficiency through technology advances. Today, the efficiency of c-Si solar cells is about 17 to 19[%] and the efficiency of PV modules is about 14 to 15 [%]. We called that electrical losses occurred in the Conversion of solar cells to PV modules are CTM loss(Cell To Module loss), the CTM loss typically has a value of about3~5[%]. The more efficiency of solar cell increase, differences are larger because the efficiency decrease owing to physical or technical problems occurred in the Conversion of solar cells to PV modules. In this study, the power loss factors occurred in the Conversion of solar cells to PV modules are analyzed and it is proposed that how to reduce losses of the PV module. The types of power loss factor are (1)losses of front glass and encapsulant(generally EVA sheet), (2)losses by sorting miss, (3)losses by interconnection, (4)losses by the field aging of PV modules. In further study, experimental and evaluation will be conducted to make demonstrate for proposed solutions.

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ANTICANCER EFFECT OF CKD-602(BELOTECAN, CAMTOBELL$^{(R)}$) ON THE ORAL CANCER CELL LINES (구강암 세포주에 대한 CKD-602의 항암 효과)

  • Yun, Pil-Young;Ok, Yong-Ju;Myoung, Hoon;Lee, Jong-Ho;Kim, Myung-Jin
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.31 no.1
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    • pp.7-12
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    • 2005
  • Purpose: CKD-602, a newly developed water-soluble campthotecin analogue, is a anticancer agent which act as a DNA topoisomerase I inhibitor. CKD-602 is known as more potent and tolerable agent. The main purposes of this study were to measure the cytotoxic effect of CKD-602 on the oral cancer cell lines and to evaluate the apoptotic aspect of dead cells. Materials and Methods: To determine the cytotoxic effect of CKD-602 on the oral cancer cell lines in comparison with various cell lines, such as lung cancer and colon cancer cell lines, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay was performed. And apoptosis was analyzed using fluorescence-activated cell sorting(FACS) system. Results: CKD-602 decreased the viability of malignant cells in a dose dependent manner and in a time dependent manner. CKD-602 showed excellent cytotoxicity to the oral cancer cell lines. Also, apoptotic portion was increased in a dose dependent manner. Conclusion: These findings indicated that CKD-602 induced apoptotic cell death in the various cell lines including oral cancer cell lines. From the results, it was suggested that CKD-602 would be a potential therapeutic agent for the oral cancer. More successive researches on the anticancer effect of CKD-602 should be performed.

Enhanced Rg3 negatively regulates Th1 cell responses

  • Cho, Minkyoung;Choi, Garam;Shim, Inbo;Chung, Yeonseok
    • Journal of Ginseng Research
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    • v.43 no.1
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    • pp.49-57
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    • 2019
  • Background: Korean Red Ginseng (KRG; Panax ginseng Meyer) is a widely used medicinal herb known to exert various immune modulatory functions. KRG and one of its purified components, ginsenoside Rg3, are known to possess anti-inflammatory activities. How they impact helper T cell-mediated responses is not fully explored. In this study, we attempted to evaluate the effect of KRG extract (KRGE) and ginsenoside Rg3 on Th1 cell responses. Methods: Using well-characterized T cell in vitro differentiation systems, we examined the effects of KRGE or enhanced Rg3 on the Th1-inducing cytokine production from dendritic cells (DC) and the naïve $CD4^+$ T cells differentiation to Th1 cells. Furthermore, we examined the change of Th1 cell population in the intestine after treatment of enhanced Rg3. The influence of KRGE or enhanced Rg3 on Th1 cell differentiation was evaluated by fluorescence-activated cell sorting, enzyme-linked immunosorbent assay, and quantitative real-time polymerase chain reaction. Results: KRGE significantly inhibited the production level of IL-12 from DCs and subsequent Th1 cell differentiation. Similarly, enhanced Rg3 significantly suppressed the expression of interferon gamma ($IFN{\gamma}$) and T-bet in T cells under Th1-skewing condition. Consistent with these effects in vitro, oral administration of enhanced Rg3 suppressed the frequency of Th1 cells in the Peyer's patch and lamina propria cells in vivo. Conclusion: Enhanced Rg3 negatively regulates the differentiation of Th1 cell in vitro and Th1 cell responses in the gut in vivo, providing fundamental basis for the use of this agent to treat Th1-related diseases.