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Development of a New Duplex Real-Time Polymerase Chain Reaction Assay for Detection of Dicer in G. gallus

  • Ji, Xiaolin;Wang, Qi;Gao, Yulong;Wang, Yongqiang;Qin, Liting;Qi, Xiaole;Gao, Honglei;Wang, Xiaomei
    • Journal of Microbiology and Biotechnology
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    • v.23 no.5
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    • pp.630-636
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    • 2013
  • Recently, there has been a growing body of evidence showing that cellular microRNAs (miRNAs) are involved in virus-host interactions. Numerous studies have focused on analyses of the expression profiles of cellular miRNAs, but the expression patterns of Dicer, which is responsible for the generation of miRNAs, have only rarely been explored in Gallus gallus. We developed a duplex real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay for the relative quantification of the mRNAs of Dicer and ${\beta}$-actin in G. gallus. To apply this method, the expression of Dicer in avian cells after infection with avian leukosis virus subgroup J (ALV-J) was detected using our established duplex real-time RT-PCR. The duplex real-time RT-PCR assay is sufficiently sensitive, specific, accurate, reproducible, and cost-effective for the detection of Dicer in G. gallus. Furthermore, this study, for the first time, demonstrated that ALV-J can induce differential expression of Dicer mRNA in the ALV-J-infected cells.

Pichia pastoris: A Recombinant Microfactory for Antibodies and Human Membrane Proteins

  • Goncalves, A.M.;Pedro, A.Q.;Maia, C.;Sousa, F.;Queiroz, J.A.;Passarinha, L.A.
    • Journal of Microbiology and Biotechnology
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    • v.23 no.5
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    • pp.587-601
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    • 2013
  • During the last few decades, it has become evident that the compatibility of the yeast biochemical environment with the ability to process and translate the RNA transcript, along with its capacity to modify a translated protein, are relevant requirements for selecting this host cell for protein expression in several pharmaceutical and clinical applications. In particular, Pichia pastoris is used as an industrial host for recombinant protein and metabolite production, showing a powerful capacity to meet required biomolecular target production levels in high-throughput assays for functional genomics and drug screening. In addition, there is a great advantage to using P. pastoris for protein secretion, even at high molecular weights, since the recovery and purification steps are simplified owing to relatively low levels of endogenous proteins in the extracellular medium. Clearly, no single microexpression system can provide all of the desired properties for human protein production. Moreover, chemical and physical bioprocess parameters, including culture medium formulation, temperature, pH, agitation, aeration rates, induction, and feeding strategies, can highly influence product yield and quality. In order to benefit from the currently available wide range of biosynthesis strategies using P. pastoris, this mini review focuses on the developments and technological fermentation achievements, providing both a comparative and an overall integration analysis. The main aim is to highlight the relevance and versatility of the P. pastoris biosystem to the design of more cost-effective microfactories to meet the increasing demands for recombinant membrane proteins and clinical antibodies for several therapeutic applications.

A Long Non-Coding RNA snaR Contributes to 5-Fluorouracil Resistance in Human Colon Cancer Cells

  • Lee, Heejin;Kim, Chongtae;Ku, Ja-Lok;Kim, Wook;Kim Yoon, Sungjoo;Kuh, Hyo-Jeong;Lee, Jeong-Hwa;Nam, Suk Woo;Lee, Eun Kyung
    • Molecules and Cells
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    • v.37 no.7
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    • pp.540-546
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    • 2014
  • Several types of genetic and epigenetic regulation have been implicated in the development of drug resistance, one significant challenge for cancer therapy. Although changes in the expression of non-coding RNA are also responsible for drug resistance, the specific identities and roles of them remain to be elucidated. Long non-coding RNAs (lncRNAs) are a type of ncRNA (> 200 nt) that influence the regulation of gene expression in various ways. In this study, we aimed to identify differentially expressed lncRNAs in 5-fluorouracil-resistant colon cancer cells. Using two pairs of 5-FU-resistant cells derived from the human colon cancer cell lines SNU-C4 and SNU-C5, we analyzed the expression of 90 lncRNAs by qPCR-based profiling and found that 19 and 23 lncRNAs were differentially expressed in SNU-C4R and SNU-C5R cells, respectively. We confirmed that snaR and BACE1AS were down-regulated in resistant cells. To further investigate the effects of snaR on cell growth, cell viability and cell cycle were analyzed after transfection of siRNAs targeting snaR. Down-regulation of snaR decreased cell death after 5-FU treatment, which indicates that snaR loss decreases in vitro sensitivity to 5-FU. Our results provide an important insight into the involvement of lncRNAs in 5-FU resistance in colon cancer cells.

Identification of a Novel Human Lysophosphatidic Acid Acyltransferase, LPAAT-theta, Which Activates mTOR Pathway

  • Tang, Wenwen;Yuan, Jian;Chen, Xinya;Gu, Xiuting;Luo, Kuntian;Li, Jie;Wan, Bo;Wang, Yingli;Yu, Long
    • BMB Reports
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    • v.39 no.5
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    • pp.626-635
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    • 2006
  • Lysophosphatidic acid acyltransferase (LPAAT) is an intrinsic membrane protein that catalyzes the synthesis of phosphatidic acid (PA) from lysophosphatidic acid (LPA). It is well known that LPAAT is involved in lipid biosynthesis, while its role in tumour progression has been of emerging interest in the last few years. To date, seven members of the LPAAT gene family have been found in human. Here we report a novel LPAAT member, designated as LPAAT-theta, which was 2728 base pairs in length and contained an open reading frame (ORF) encoding 434 amino acids. The LPAAT-theta gene consisted of 12 exons and 11 introns, and mapped to chromosome 4q21.23. LPAAT-theta was ubiquitously expressed in 18 human tissues by RT-PCR analysis. Subcellular localization of LPAAT-theta-EGFP fusion protein revealed that LPAAT-theta was distributed primarily in the endoplasmic reticulum (ER) of COS-7 cells. Furthermore, we found that the overexpression of LPAAT-theta can induce mTOR-dependent p70S6K phosphorylation on Thr389 and 4EBP1 phosphorylation on Ser65 in HEK293T cells.

Congenital muscular dystrophy type 1A with residual merosin expression

  • Kim, Hyo Jeong;Choi, Young-Chul;Park, Hyung Jun;Lee, Young-Mock;Kim, Heung Dong;Lee, Joon Soo;Kang, Hoon-Chul
    • Clinical and Experimental Pediatrics
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    • v.57 no.3
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    • pp.149-152
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    • 2014
  • Congenital muscular dystrophy type 1A (MDC1A) is an autosomal recessive disorder characterized by hypotonia, elevated serum creatine kinase level, delayed motor milestones, white matter changes observed by brain magnetic resonance imaging, and normal intelligence. A mutation in the laminin ${\alpha}2$ (LAMA2) gene, located at 6q22-23, is a genetic cause of MDC1A. Patients have merosin (laminin ${\alpha}2$)-deficient skeletal muscles. However, the degree of merosin expression ranges from total absence to partial reduction. Patients with residual merosin expression have more variable and milder phenotypes than those with absolute merosin deficiency. We observed a Korean girl with MDC1A with residual merosin expression. Clinical presentation of this patient was typical except for late onset of the disease and external capsule involvement. Immunohistochemical staining of muscle fibers including merosin, is important to evaluate patients with hypotonia, delayed motor development, and abnormal white matter changes.

Maximizing the Selection Response by Optimal Quantitative Trait Loci Selection and Control of Inbreeding in a Population with Different Lifetimes between Sires and Dams

  • Tang, G.Q.;Li, X.W.;Zhu, L.;Shuai, S.R.;Bai, L.
    • Asian-Australasian Journal of Animal Sciences
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    • v.21 no.11
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    • pp.1559-1571
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    • 2008
  • A rule was developed to constrain the annual rate of inbreeding to a predefined value in a population with different lifetimes between sires and dams, and to maximize the selection response over generations. This rule considers that the animals in a population should be divided into sex-age classes based on the theory of gene flow, and restricts the increase of average inbreeding coefficient for new offspring by limiting the increase of the mean additive genetic relationship for parents selected. The optimization problem of this rule was formulated as a quadratic programming problem. Inputs for the rule were the BLUP estimated breeding values, the additive genetic relationship matrix of all animals, and the long-term contributions of sex-age classes. Outputs were optimal number and contributions of selected animals. In addition, this rule was combined with the optimization of emphasis given to QTL, and further increased the genetic gain over the planning horizon. Stochastic simulations of closed nucleus schemes for pigs were used to investigate the potential advantages obtained from this rule by combining the standard QTL selection, optimal QTL selection and conventional BLUP selection. Results showed that the predefined rates of inbreeding were actually achieved by this rule in three selection strategies. The rule obtained up to 9.23% extra genetic gain over truncation selection at the same rates of inbreeding. The combination of the extended rule and the optimization of emphasis given to QTL allowed substantial increases in selection response at a fixed annual rate of inbreeding, and solved substantially the conflict between short-term and long-term selection response in QTL-assisted selection schemes.

Proteome Analysis of Mouse Adipose Tissue and Colon Tissue using a Novel Integrated Data Processing Pipeline

  • Park, Jong-Moon;Han, Na-Young;Kim, Hokeun;Hwang, Injae;Kim, Jae Bum;Hahm, Ki-Baik;Lee, Sang-Won;Lee, Hookeun
    • Mass Spectrometry Letters
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    • v.5 no.1
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    • pp.16-23
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    • 2014
  • Liquid chromatography based mass spectrometry (LC-MS) is a key technology for analyzing highly complex and dynamic proteome samples. With highly accurate and sensitive LC-MS analysis of complex proteome samples, efficient data processing is another critical issue to obtain more information from LC-MS data. A typical proteomic data processing starts with protein database search engine which assigns peptide sequences to MS/MS spectra and finds proteins. Although several search engines, such as SEQUEST and MASCOT, have been widely used, there is no unique standard way to interpret MS/MS spectra of peptides. Each search engine has pros and cons depending on types of mass spectrometers and physicochemical properties of peptides. In this study, we describe a novel data process pipeline which identifies more peptides and proteins by correcting precursor ion mass numbers and unifying multi search engines results. The pipeline utilizes two open-source software, iPE-MMR for mass number correction, and iProphet to combine several search results. The integrated pipeline identified 25% more proteins in mouse epididymal adipose tissue compared with the conventional method. Also the pipeline was validated using control and colitis induced colon tissue. The results of the present study shows that the integrated pipeline can efficiently identify increased number of proteins compared to the conventional method which can be a breakthrough in identification of a potential biomarker candidate.

Performance Evaluation of a Differentiated Service Mechanism by Traffic Models and Weight Factor (트래픽 모델과 Weight Factor에 의한 차등 서비스 메커니즘의 성능평가)

  • 전용희;박수영
    • The Journal of Korean Institute of Communications and Information Sciences
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    • v.26 no.11C
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    • pp.10-23
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    • 2001
  • It is difficult to satisfy the QoS(Quality of Service) guarantee which is required by real-time application services only with the Best-Effort(BE) service adopted in the current Internet. Therefore, worldwide research is being made on the method of QoS provisioning. Among them, the QoS guarantee mechanism using the Diffserv(Differentiated Service) was discussed in this paper. First we analyzed how the DiffServ performance was affected by traffic models. For this, we performed the research for the random, bursts, and self-similar traffic modeling method. We then designed and implemented an OPNET simulator, and performed the simulation 7d performance evaluation for diverse input parameters. Based on the results of performance evaluation, it was confirmed that QoS guarantee is possible for the EF(Expedited Forwarding) class with the DiffServ function under every environments considered and the service separation between EF and BE(Best Effort) classes is also possible. We also analyzed the performance variation and dynamic behavior of DiffServ mechanism based on the resource allocation between E? and BE classes in WFQ(Weighted Fair Queueing).

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Effects of the $V_2$$O_5$ Additive on ${ZnNb_2}{O_6}$ Microwave Dielectrics

  • Yoo, Sang-Im;Kim, Dong-Wan;Wee, Sung-Hun;Hong, Kug-Sun
    • Journal of the Korean Ceramic Society
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    • v.38 no.4
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    • pp.308-313
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    • 2001
  • We report the effects of the V$_2$O$_{5}$ additive on the sintering behavior and microwave dielectric properties of ZnNb$_2$O$_{6}$ ceramics. Densification temperatures of V$_2$O$_{5}$-doped ZnNb$_2$O$_{6}$ samples are lowered to the range of 875-9$25^{\circ}C$ because of the liquid phase sintering. Doped samples are composed of a Zn(Nb,V)$_2$O$_{6}$ solid solution and second phases. Up to 5 wt% V$_2$O$_{5}$ is the only second phase, however, V$_2$O$_{5}$ also exists as the second phase for 10 wt% V$_2$O$_{5}$ addition. In comparison with reported values of undoped ZnNb$_2$O$_{6}$ ceramics, microwave properties of V$_2$O$_{5}$-doped ZnNb$_2$O$_{6}$ samples are seriously degraded, which is confirmed to originate from the second phases. The optimum microwave properties (Q$\times$f=13,800, $\varepsilon$$_{r}$=23, $\tau$$_{f}$=-66ppm/$^{\circ}C$) are obtained from ZnNb$_2$O$_{6}$ with the addition of 5 wt% V$_2$O$_{5}$ sintered at 90$0^{\circ}C$.90$0^{\circ}C$.EX>.

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Effects of Aromatherapy on Sleep Quality: A Systematic Review and Meta-Analysis (아로마테라피가 수면에 미치는 효과: 체계적 문헌고찰 및 메타분석)

  • Kim, Mi-Eun;Jun, Ji Hee;Hur, Muyng-Haeng
    • Journal of Korean Academy of Nursing
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    • v.49 no.6
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    • pp.655-676
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    • 2019
  • Purpose: The purpose of this study was to investigate the effects of aromatherapy on sleep quality. Methods: This is a systematic review of randomized controlled trial studies (PROSPERO registration number CRD42017064519). In this study, the PICO were adults and the elderly, aromatherapy intervention, a comparative intervention with the control and placebo oil groups, and sleep. The selected articles were in English, Korean, and Chinese. Results: The results of the meta-analysis showed that the effect sizes of the experimental group were 1.03 (n=763, SMD=1.03, 95% CI 0.66 to 1.39) (Z=5.47, p<.001). In the aromatherapy intervention group, the effect size of sleep was statistically significant (QB=9.39, df=2, p=.009), with a difference of 0.77 for inhalation, 1.12 for oral intake and 2.05 for massage. A post-analysis showed that the effect of massage on sleep was significantly greater than the inhalation method. The regression coefficient of the intervention period, B=0.01 (Z=1.43, p=.154), also showed that the longer the intervention period, the larger the effect size; however, it was not statistically significant. Conclusion: A total of 23 literature analyses showed that aromatherapy is effective in improving quality of sleep, and the massage method is more effective in improving quality of sleep than the inhalation method. A meta-ANOVA showed that the aromatherapy intervention affected the high heterogeneity of the effect size. Thus, future research with stricter control in methods and experimental procedures is necessary.