• Asian Pacific Journal of Cancer Prevention
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• 제15권23호
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• pp.10181-10185
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• 2015

Background: MicroRNAs, small noncoding RNA molecules, can regulate mammalian cell growth, apoptosis and differentiation by controlling the expression of target genes. The aim of this study was to investigate the function of miR-323-5p in the glioma cell line, U251. Materials and Methods: After over-expression of miR-323-5p using miR-323-5p mimics, cell growth, apoptosis and migration were tested by MTT, flow cytometry and cell wound healing assay, respectively. We also assessed the influence of miR-323-5p on the mRNA expression of IGF-1R by quantitative real-time reverse transcriptase PCR (qRT-PCR), and on the protein levels by Western blot analysi. In addition, dual-luciferase reporter assays were performed to determine the target site of miR-323-5p to IGF-1R 3'UTR. Results: Our findings showed that over-expression of miR-323-5p could promote apoptosis of U251 and inhibit the proliferation and migration of the glioma cells. Conclusions: This study demonstrated that increased expression of miR-323-5p might be related to glioma progression, which indicates a potential role of miR-323-5p for clinical therapy.

• Asian Pacific Journal of Cancer Prevention
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• 제15권15호
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• pp.6269-6273
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• 2014

Growing evidence suggests that miR-150-5p has an important role in regulating genesis of various types of cancer. However, the roles and the underlying mechanisms of miR-150-5p in development of colorectal cancer (CRC) remain largely unknown. Transwell chambers were used to analyze effects on cell migration and invasion by miR-150-5p. Quantitative real-time PCR (qRT-PCR), Western blotting and dual-luciferase 3' UTR reporter assay were carried out to identify the target genes of miR-150-5p. In our research, miR-150-5p suppressed CRC cell migration and invasion, and MUC4 was identified as a direct target gene. Its effects were partly blocked by re-expression of MUC4. In conclusiomn, miR-150-5p may suppress CRC metastasis through directly targeting MUC4, highlighting its potential as a novel agent for the treatment of CRC metastasis.

• Journal of Microbiology and Biotechnology
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• 제22권4호
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• pp.479-483
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• 2012

The secretory efficiency of recombinant xylanase xynB from yeast Pichia pastoris between the ${\alpha}$-factor preprosequence and a classical mammalian signal peptide derived from bovine ${\beta}$-casein was compared. The results showed that although the bovine ${\beta}$-casein signal peptide could direct high-level secretion of recombinant xylanase, it was relatively less efficient than the ${\alpha}$-factor preprosequence. In contrast, the bovine ${\beta}$-casein signal peptide caused remarkably more recombinant xylanase trapped intracellularly. Real-time RT-PCR analysis indicated that the difference in the secretory level between the two signal sequences was not due to the difference in the transcriptional efficiency.

• 환경생물
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• 제37권1호
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• pp.72-81
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• 2019

Bisphenol A (BPA)는 대표적인 내분비계장애물질로 수서생물의 성장, 발생, 그리고 생식에 유해한 영향을 주는 것으로 알려져 있다. 본 연구는 기수산 물벼룩(Diaphanosoma celebensis)에서 48시간 BPA 노출 후 산화적 스트레스 반응을 조사하기 위하여 qRT-PCR을 이용한 항산화 유전자 발현 변화, 항산화 효소 활성, 총 단백질 함량 분석을 수행하였다. $3mg\;L^{-1}$의 BPA에 48시간 노출된 D. celebensis에서 모든 항산화 유전자(Cu/Zn-SOD, Mn-SOD, CAT)의 발현량이 유의하게 증가하였다. 특히 세 종류의 GST isoforms (GST-kappa, GST-mu, GST-theta)는 가장 낮은 농도인 $0.12mg\;L^{-1}$ BPA에 48시간 노출된 실험군에서도 유의하게 증가하였으며, GST-mu의 발현양이 상대적으로 가장 높게 나타났다. SOD 활성은 BPA 농도에 의존적으로 유의하게 증가하였으며, 총 단백질 함량은 BPA 노출에 대해 감소되는 양상을 보였다. 이러한 결과는 BPA가 D. celebensis에서 산화적 스트레스를 유발하였고, 이 과정에서 이들 항산화 유전자가 생물방어기전으로 참여한다는 것을 의미한다. 본 연구는 BPA가 해양생물에 미치는 영향에 대한 분자적 기전을 이해하는 데 도움이 될 것이다.

• Mycobiology
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• 제47권1호
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• pp.59-65
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• 2019

Agrobacterium tumefaciens-mediated transformation (ATMT), as a simple and versatile method, achieves successful transformation in the yeast-like cells (YLCs) of Tremella fuciformis with lower efficiency. Establishment of a more efficient transformation system of YLCs is important for functional genomics research and biotechnological application. In this study, an enzymolysis-assisted ATMT method was developed. The degradation degree of YLCs depends on the concentration and digestion time of Lywallzyme. Lower concentration (${\leq}0.1%$) of Lywallzyme was capable of formation of limited wounds on the surface of YLCs and has less influence on their growth. In addition, there is no significant difference of YLCs growth among groups treated with 0.1% Lywallzyme for different time. The binary vector pGEH under the control of T. fuciformis glyceraldehyde-3-phosphate dehydrogenase gene (gpd) promoter was utilized to transform the enzymolytic wounded YLCs with different concentrations and digestion time. The results of PCR, Southern blot, quantitative real-time PCR (qRT-PCR) and fluorescence microscopy revealed that the T-DNA was integrated into the YLCs genome, suggesting an efficient enzymolysis-assisted ATMT method of YLCs was established. The highest transformation frequency reached 1200 transformants per $10^6$ YLCs by 0.05% (w/v) Lywallzyme digestion for 15 min, and the transformants were genetically stable. Compared with the mechanical wounding methods, enzymolytic wounding is thought to be a tender, safer and more effective method.

• Asian-Australasian Journal of Animal Sciences
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• 제31권10호
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• pp.1550-1557
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• 2018

Objective: Circular RNAs (circRNAs) are a newfound class of non-coding RNA in animals and plants. Recent studies have revealed that circRNAs play important roles in cell proliferation, differentiation, autophagy and apoptosis during development. However, there are few reports about muscle development-related circRNAs in livestock. Methods: RNA sequencing analysis was employed to identify and annotate circRNAs from longissimus dorsi of sheep. Reverse transcription followed by real-time quantitative (q) polymerase chain reaction (PCR) analysis verified the presence of these circRNAs. Targetscan7.0 and miRanda were used to analyse the interaction of circRNA-microRNA (miRNA). To investigate the function of circRNAs, an experiment was conducted to perform enrichment analysis hosting genes of circRNAs using gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathways. Results: About 75.5 million sequences were obtained from RNA libraries of sheep skeletal muscle. These sequences were mapped to 729 genes in the sheep reference genome. We identified 886 circRNAs, including numerous circular intronic RNAs and exonic circRNAs. Reverse transcription PCR (RT-PCR) and DNA sequencing analysis confirmed the presence of several circRNAs. Real-Time RT-PCR analysis exhibited resistance of sheep circRNAs to RNase R digestion. We found that many circRNAs interacted with muscle-specific miRNAs involved in growth and development of muscle, especially circ776. The GO and KEGG enrichment analysis showed that hosting genes of circRNAs was involved in muscle cell development and signaling pathway. Conclusion: The study provides comprehensive expression profiles of circRNAs in sheep skeletal muscle. Our study offers a large number of circRNAs to facilitate a better understanding of their roles in muscle growth. Meanwhile, we suggested that circ776 could be analyzed in future study.

• 한국해양생명과학회지
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• 제7권2호
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• pp.145-153
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• 2022

수은은 생물 축적과 먹이사슬을 통한 생물 농축되며, 미량에서도 유해한 영향을 나타내기 때문에 해양 환경 내에서 중요한 문제가 되고 있다. 그러나 해양 소형 갑각류에 대한 수은의 생물 영향은 다른 금속에 비해 연구가 미흡하다. 본 연구에서는 기수산 물벼룩 Diaphanosoma celebensis을 아치사 농도(0.2, 0.4, 0.8 ㎍/l)의 무기 수은(HgCl₂)에 48시간 노출시킨 후, 대사 관련 유전자의 발현 양상을 조사하였다. 해독효소 유전자 5종(cytochrome P450; cyp360A1, cyp361A1, cyp4AP3, cyp4C122, cyp370C5)과 소화효소 6종(alpha amylase (AMY), alpha amylase related protein (AMY-like), trypsin (TRYP), chymotrypsin-like protein (CHY), lipase (LIP), pancreatic lipase-related protein (PLRP))의 유전자 발현을 quantitative real time reverse transcription polymerase chain reaction (qRT-PCR)을 이용하여 분석하였다. Cyp 유전자의 경우 clan2에 속하는 cyp370C5와 clan4에 속하는 cyp4AP3 유전자의 발현이 농도 의존적으로 유의하게 증가하였다. 한편 소화효소 유전자 중에서는 단백질 소화와 관련 있는 TRYP 유전자의 발현이 농도 의존적으로 증가하였다. 이러한 결과는 cyp370C5와 cyp4AP3가 수은 독성을 해독하는 과정에서 중요한 역할을 담당할 것으로 보이며, 수은이 소화효소 유전자의 발현을 조절함으로써 에너지 대사에 영향을 미칠 수 있음을 제시한다. 본 연구는 해양 소형 갑각류에서 수은에 대한 분자 수준의 영향을 이해하는데 도움이 될 것이다.

• Journal of Animal Science and Technology
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• 제57권8호
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• pp.28.1-28.7
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• 2015

Background: The physiologic characteristics of the cashmere trait and many of the differentially expressed genes relevant to hair cycling have been extensively studied, whereas genes involved in the prenatal development of hair follicles have been poorly investigated in cashmere goats. The aim of this study, therefore, was to quantify the time-course changes in the expressions of $TR{\alpha}$ and CRABPII genes in the fetal skin of Chinese cashmere goats at the multiple embryonic days (E70, E75, E80, E90, E100, E120 and E130) using real-time quantitative PCR (RT-qPCR). Results: RT-qPCR showed that $TR{\alpha}$ was expressed at E70 with relatively high level and then slightly decreased (E75, E80, and E90). The highest expression of $TR{\alpha}$ mRNA was revealed at E130 (P > 0.05). The expression pattern of CRABPII mRNA showed an 'up-down-up' trend, which revealed a significantly highest expression at E75 (P < 0.05) and was down-regulated during E80 to E120 (P < 0.05) and mildly increased at E130, subsequently. Conclusion: This study demonstrated that $TR{\alpha}$ and CRABPII genes expressed in different levels during prenatal development of cashmere. The present study will be helpful to provide the comprehensive understanding of $TR{\alpha}$ and CRABPII genes expressions during cashmere formation and lay the ground for further studies on their roles in regulation of cashmere growth in goats.

• Asian Pacific Journal of Cancer Prevention
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• 제14권11호
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• pp.6281-6286
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• 2013

Background: To further investigate the molecular basis of lung cancer development, we utilize a microarray to identify differentially expressed genes associated with various TNM stages of adenocarcinoma, a subtype with increasing incidence in recent years in China. Methods: A 35K oligo gene array, covering about 25,100 genes, was used to screen differentially expressed genes among 90 tumor samples of lung adenocarcinoma in various TNM stages. To verify the gene array data, three genes (Zimp7, GINS2 and NAG-1) were confirmed by real-time RT-PCR in a different set of samples from the gene array. Results: First, we obtained 640 differentially expressed genes in lung adenocarcinomas compared to the surrounding normal lung tissues. Then, from the 640 candidates we identified 10 differentially expressed genes among different TNM stages (Stage I, II and IIIA), of which Zimp7, GINS2 and NAG-1 genes were first reported to be present at a high level in lung adenocarcinoma. The results of qRT-PCR for the three genes were consistent with those from the gene array. Conclusions: We identified 10 candidate genes associated with different TNM stages in lung adenocarcinoma in the Chinese population, which should provide new insights into the molecular basis underlying the development of lung adenocarcinoma and may offer new targets for the diagnosis, therapy and prognosis prediction.

• Biomolecules & Therapeutics
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• 제24권1호
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• pp.19-24
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• 2016

Acute myocardial infarction (AMI) remains a leading cause of morbidity and mortality worldwide. The exploration of new biomarkers with high sensitivity and specificity for early diagnosis of AMI therefore becomes one of the primary task. In the current study, we aim to detect whether there is any heart specific long noncoding RNA (lncRNA) releasing into the circulation during AMI, and explore its function in the neonatal rat cardiac myocytes injury induced by $H_2O_2$. Our results revealed that the cardiac-specific lncRNA MHRT (Myosin Heavy Chain Associated RNA Transcripts) was significantly elevated in the blood from AMI patients compared with the healthy control ($^*p<0.05$). Using an in vitro neonatal rat cardiac myocytes injury model, we demonstrated that lncRNA MHRT was upregulated in the cardiac myocytes after treatment with hydrogen peroxide ($H_2O_2$) via real-time RT-PCR (qRT-PCR). Furthermore, we knockdowned the MHRT gene by siRNA to confirm its roles in the $H_2O_2$-induced cardiac cell apoptosis, and found that knockdown of MHRT led to significant more apoptotic cells than the non-target control ($^{**}p<0.01$), indicating that the lncRNA MHRT is a protective factor for cardiomyocyte and the plasma concentration of MHRT may serve as a biomarker for myocardial infarction diagnosis in humans AMI.