• Title/Summary/Keyword: RD-PCR

Search Result 94, Processing Time 0.029 seconds

고려인삼으로부터 Squalene Synthase 유전자의 Cloning 및 형질전환체 특성

  • Sim, Ju-Seon;Son, Hwa;Yang, Deok-Chun
    • Proceedings of the Ginseng society Conference
    • /
    • 2004.12a
    • /
    • pp.50-52
    • /
    • 2004
  • Introduce of gene connected with disease and transformation system of ginseng, Squalene systhase(PSS) gene cloned from and disease resistant gene were carried out for expression and transformation of plant using Agrobacterium. PSS of 35S-35S-AMV-PSS-Tnos, has been constructed which were mobilized into Agrobacterium tumefaciens strain MP 90 disarmed Ti-plasmid. PSS gene were introduced into the binary vector pRD 400. The transgenic ginseg plants were propagated using repetitive secondary embryogenesis and introduced NPTII and PSS genes of the transgenic ginseng were successfully indentified by the PCR and survival test on the medium.

  • PDF

Changes in mRNA Expression of Obesity-related Genes by GyeongshinhaeGihwan 1 (GGT1) in hGHTg (human growth hormone transgenic) obese Female Rats (암컷 hGHTg 비만 쥐에서 경신해지환(輕身解脂丸) (GGT1)에 의한 비만관련 유전자 mRNA 발현의 변화)

  • Yoon, Ki-Hyeon;Yoon, Mi-Chung;Kim, Hoon;Shin, Soon-Shik
    • Journal of Physiology & Pathology in Korean Medicine
    • /
    • v.20 no.2
    • /
    • pp.383-387
    • /
    • 2006
  • To investigate the effect of GyeongshinhaeGihwan 1(GGT1) frequently used as an anti-obesity herbal medicine in oriental medicine on the expression of obesity-related genes, we measured the changes in mRNA levels of these genes by GGT1 in human growth hormone transgenic (hGHTg) obese female rats, and these effects by GGT1 were compared with those of reductil (RD), an anti-obesity drug approved by FDA. Rats received once daily oral administrations of autoclaved water, RD, or GGT1 for 8 weeks. At the end of study, rats were sacrificed and tissues were harvested. Total RNA from adipose tissue, liver and kidney was prepared and the mRNA levels for LPL (lipoprotein lipase), $PPAR{\gamma}$ (peroxisome proliferator activated receptor-gamma), $PPAR{\delta}$ (peroxisome proliferator activated receptor-delta), leptin, $TNF{\alpha}$ (tumor necrosis factor-alpha), and internal standard G3PDH (glyceraldehyde-3-phosphate dehydrogenase) were analyzed by RT-PCR. Compared with control group, $PPAR{\gamma}$ mRNA levels of liver and kidney were decreased in both RD and GGT1 groups, and the effects were more prominent in GGT1 group than in RD group, suggesting that GGT1 is effective in the inhibition of lipid storage by decreasing the $PPAR{\gamma}$ expression. $PPAR{\delta}$ mRNA levels of adipose tissue were increased by RD and GGT1 compared with DW, and the magnitude of increase were higher in GGT1 group than in RD group, indicating that GGT1 stimulates fatty acid oxidation and energy metabolism by activating $PPAR{\delta}$ expression. GGT1 group had higher concentrations of serum leptin, a well-known inhibitor of appetite, than control and RD groups. However, The mRNA levels of leptin, LPL, and $TNF{\alpha}$ were not changed by GGT1. These results indicate that GGT1 can prevent obesity in hGHTg obese female rats by down-regulating and up-regulating the mRNA expression of $PPAR{\gamma}$ and $PPAR{\delta}$, respectively, and that this anti-obesity effects were more pronounced in GGT1 group compared with RD group. In addition, GGT1 seems to inhibit obesity by increasing the circulating leptin levels.

Effects of GyeongshinhaeGihwan 1(GGT1) on the Expression of Obesity-related Genes in Obese Male hGHTg Rats (경신해지환(輕身解脂丸) (GGT1)이 형질전환 비만모델 hGHTg 수컷 쥐의 비만관련 유전자 발현에 미치는 영향)

  • Jung Yang-Sam;Yoon Mi-Chung;Kim Gyeong-Cheol;Shin Soon-Shik
    • Journal of Physiology & Pathology in Korean Medicine
    • /
    • v.20 no.1
    • /
    • pp.93-97
    • /
    • 2006
  • To investigate whether GyeongshinhaeGihwan 1(GGT1), an anti-obesity herbal medicine widely used in oriental medicine, regulates the expression of obesity-related genes, we measured the changes in mRNA levels of these genes by GGT1 in human growth hormone transgenic (hGHTg) obese male rats, and these effects by GGT1 were compared with those of reductil (RD), an anti-obesity drug approved by FDA. Rats received once daily oral administrations of autoclaved water, RD, or GGT1 for 8 weeks. At the end of study, rats were sacrificed and tissues were harvested. Total RNA from adipose tissue, liver and kidney was prepared and the mRNA levels for LPL (lipoprotein lipase), PPAR $\gamma$ (peroxisome proliferator activated receptor-gamma), PPAR$\delta$ (peroxisome proliferator activated receptor-delta), leptin, TNF$\alpha$ (tumor necrosis factor-alpha), and internal standard G3PDH (glyceraldehyde-3- phosphate dehydrogenase) were analyzed by RT-PCR. PPAR$\gamma$ mRNA levels of liver and kidney were decreased in drug-treated groups compared with control group and the decrease of PPAR$\gamma$ expression was more prominent in GGT1 group than in RD group, suggesting that GGT1 is effective in the inhibition of adipogenesis and lipid storage by decreasing the PPAR$\gamma$ expression. In contrast, PPAR$\delta$ mRNA levels of adipose tissue and kidney were increased by RD and GGT1 , and the magnitudes of increase were higher in GGT1 group than in RD group, indicating that GGT1 stimulates fatty acid oxidation and energy metabolism by activating PPAR$\delta$ expression, Compared with control and RD groups, GGT1 group had higher concentrations of serum leptin, a well-known inhibitor of appetite. However, The mRNA levels of leptin, LPL, and TNF$\alpha$ were not changed by GGT1 and RD, compared with DW. These results demonstrate that GGT1 not only decreases PPAR$\gamma$ expression of liver and kidney, but also increases PPAR$\delta$ expression of adipose tissue and kidney, leading to the regulation of obesity and that these effects were more pronounced in GGT1 group compared with RD group. In addition, GGT1 seems to prevent obesity by increasing the serum leptin levels.

Association Between Polymorphisms of XRCC1 Arg399Gln and XPD Lys751Gln Genes and Prognosis of Colorectal Cancer in a Chinese Population

  • Gan, Yi;Li, Xiao-Rong;Chen, Dao-Jin;Wu, Jun-Hui
    • Asian Pacific Journal of Cancer Prevention
    • /
    • v.13 no.11
    • /
    • pp.5721-5724
    • /
    • 2012
  • We conducted this study to detect associations between XRCC1 Arg399Gln and XPD Lys751Gln genotypes and survival of colorectal cancer patients treated with 5-FU/oxalipatin chemotherapy. We included 289 Chinese patients with advanced colorectal cancer, who had received 5-FU/oxalipatin chemotherapy as first-line treatment from January 2005 to January 2007. All patients were followed up till Nov. 2011. Genotyping for XRCC1 Arg399Gln and XPD Lys751Gln polymorphisms was based upon duplex polymerase-chain-reaction with the PCR-RFLP method. In our study, we found the XRCC1 399 Gln/Gln genotype to confer significantly higher rates of response to chemotherapy when compared to the Arg/Arg genotype [OR (95% CI)= 2.56(1.57-2.55)]. patients with the XPD 751 Gln/Gln genotype had significantly higher rates of response to chemotherapy [OR (95% CI)= 1.54(0.87-2.65)] and those with the XRCC1 399 Gln/Gln genotype had a longer average survival time and significantly lower risk of death than did those with the Arg/Arg genotype [HR (95% CI)= 0.66(0.36-0.95)]. Similarly, those carrying the XPD 751Gln/Gln genotype had 0.51-fold the risk of death of those with XPD 751Lys/Lys [HR (95% CI)= 0.51(0.33 -0.94)]. In conclusion, it is suggested that the XRCC1 Arg399Gln and XPD Lys751Gln polymorphisms should be routinely assessed to determine colorectal patients who are more likely to benefit from 5-FU/oxalipatin chemotherapy.

Antagonistic Regulation of Arabidopsis Growth by Brassinosteroids and Abiotic Stresses

  • Chung, Yuhee;Kwon, Soon Il;Choe, Sunghwa
    • Molecules and Cells
    • /
    • v.37 no.11
    • /
    • pp.795-803
    • /
    • 2014
  • To withstand ever-changing environmental stresses, plants are equipped with phytohormone-mediated stress resistance mechanisms. Salt stress triggers abscisic acid (ABA) signaling, which enhances stress tolerance at the expense of growth. ABA is thought to inhibit the action of growth-promoting hormones, including brassinosteroids (BRs). However, the regulatory mechanisms that coordinate ABA and BR activity remain to be discovered. We noticed that ABA-treated seedlings exhibited small, round leaves and short roots, a phenotype that is characteristic of the BR signaling mutant, brassinosteroid insensitive1-9 (bri1-9). To identify genes that are antagonistically regulated by ABA and BRs, we examined published Arabidopsis microarray data sets. Of the list of genes identified, those upregulated by ABA but downregulated by BRs were enriched with a BRRE motif in their promoter sequences. After validating the microarray data using quantitative RT-PCR, we focused on RD26, which is induced by salt stress. Histochemical analysis of transgenic Arabidopsis plants expressing RD26pro:GUS revealed that the induction of GUS expression after NaCl treatment was suppressed by co-treatment with BRs, but enhanced by co-treatment with propiconazole, a BR biosynthetic inhibitor. Similarly, treatment with bikinin, an inhibitor of BIN2 kinase, not only inhibited RD26 expression, but also reduced the survival rate of the plant following exposure to salt stress. Our results suggest that ABA and BRs act antagonistically on their target genes at or after the BIN2 step in BR signaling pathways, and suggest a mechanism by which plants fine-tune their growth, particularly when stress responses and growth compete for resources.

Acibenzolar-S-Methyl(ASM)-Induced Resistance against Tobamoviruses Involves Induction of RNA-Dependent RNA Polymerase(RdRp) and Alternative Oxidase(AOX) Genes

  • Madhusudhan, Kallahally Nagendra;Deepak, Saligrama Adavigowda;Prakash, Harishchandra Sripathi;Agrawal, Ganesh Kumar;Jwa, Nam-Soo;Rakwal, Randeep
    • Journal of Crop Science and Biotechnology
    • /
    • v.11 no.2
    • /
    • pp.127-134
    • /
    • 2008
  • Tobamoviruses are the major viral pathogens of tomato and bell pepper. The preliminary results showed that Acibenzolar-Smethyl(ASM; S-methylbenzo(1,2,3) thiadiazole-7-carbothiate) pre-treatment to tomato and tobacco plants reduces the concentration of Tomato mosaic tobamovirus(ToMV) and Tobacco mosaic tobamovirus(TMV) in tomato and bell pepper seedlings, respectively. Pre-treatment of the indicator plant(Nicotiana glutinosa) with the ASM followed by challenge inoculation with tobamoviruses produced a reduced number and size of local lesions(67 and 79% protection over control to TMV and ToMV inoculation, respectively). In order to understand the mechanism of resistance the gene expression profiles of antiviral genes was examined. RT-PCR products showed higher expression of two viral resistance genes viz., alternative oxidase(AOX) and RNA dependent RNA polymerase(RdRp) in the upper leaves of the ASM-treated tomato plants challenge inoculation with ToMV. Further, the viral concentration was also quantified in the upper leaves by reverse transcription PCR using specific primer for movement protein of ToMV, as well as ELISA by using antisera against tobamoviruses. The results provided additional evidence that ASM pre-treatment reduced the viral movement to upper leaves. The results suggest that expressions of viral resistance genes in the host are the key component in the resistance against ToMV in the inducer-treated tomato plants.

  • PDF

Pulsed-Field Gel Electrophoresis and Mutation Typing of gyrA Gene of Quinolone-Resistant Salmonella enterica Serovar Paratyphi A Isolated from Outbreak and Sporadic Cases,1998-2002, Korea

  • KIM SHUKHO;OK YOUNG LIM;SEONG HAN KIM;JUN YOUNG KIM;YEON HO KANG;BOK KWON LEE
    • Journal of Microbiology and Biotechnology
    • /
    • v.13 no.1
    • /
    • pp.155-158
    • /
    • 2003
  • In early 2002, over 200 people in the city of Pusan. Korea suffered from paratyphoid fever resulting from Salmonella Paratyphi A Infection. Antimicrobial susceptibility tests and Xbal pulsed-field gel electrophoresis (PFCE) were conducted to 54 Salmonella Paratyphi A isolated from humans during the period of 1998 to 2002. Most of the isolates ($83\%$) were only nalidixic acid-resistant and $78\%$ were X 1 PFGE patterns. Also, we measured the MIC of ciprofloxacin and screened gyrA mutation(5) using allele- specific PCR and restriction fragment length polymorphism (AS-PCR-RFLP). The representative 5 isolates in 2002 and 1 isolate in 2000 were $1{\mu}g/ml$ of MIC and had mutation at the 83rd codon in gyrA. These data suggest that the outbreak in the early 2002 might have been due to dissemination of the strain present In 2000. Also, decreased susceptibility to ciprofloxacin was partly due to the mutation at the 83rd codon in gyrA.

EXPRESSION OF TGF-$\beta$ AND IGF-I DURING OSSEOINTEGRATION OF TITANIUM IMPLANT (타이타늄 임프란트 골유착시 TGF-$\beta$와 IGF-I의 발현)

  • Lee, In-Woong;Song, Hyun-Chul;Jee, Yu-Jin
    • Maxillofacial Plastic and Reconstructive Surgery
    • /
    • v.27 no.2
    • /
    • pp.123-130
    • /
    • 2005
  • Many of the molecular and genotypic events taking place at the osteoblast cell level during bone-implant integration are still largely unknown. The objective of this study was to examine expression patterns of TGF-$\beta$ and IGF-I related genes during bone-implant integration. Titanium implants with machined surface were placed into 8 rabbit tibias. At 3rd, 7th, 14th, 28th day after implantation, the expression pattern of TGF-$\beta$ and IGF-I genes in bone with or without implant was examined using reverse transcriptase-polymerase chain reaction (RT-PCR). At the same time, histomorphometric analysis was evaluated, respectively. The bone-to-implant contacts (BIC) of experimental groups were 5.2%, 6.2%, 6.6%, 24.6% at 3rd, 7th, 14th, 28th day. This indicated that newly formed bone increased at the implant surface in bone marrow space after implantation. The expressions of TGF-$\beta$ and IGF-I were higher in implantation groups than untreated control groups during all experimental days. The increased expression of TGF-$\beta$ and IGF-I genes may be associated with the increased bone-to-implant contact. This result provided the evidence for existing biologic differences in tissue response after implantation and helped us to understand molecular biologic processes in tissue-implant integration.

Comparison of clinical diagnostic performance between commercial RRT-LAMP and RT-qPCR assays for SARS-CoV-2 detection

  • Kim, Hye-Ryung;Park, Jonghyun;Han, Hyung-Soo;Kim, Yu-Kyung;Jeon, Hyo-Sung;Park, Seung-Chun;Park, Choi-Kyu
    • Korean Journal of Veterinary Service
    • /
    • v.44 no.3
    • /
    • pp.163-168
    • /
    • 2021
  • The rapid and reliable detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays a key role in isolating infected patients and preventing further viral transmission. In this study, we evaluated the clinical diagnostic performances of a commercial real-time reverse transcription loop-mediated isothermal amplification (RRT-LAMP) assay (Isopollo® COVID-2 assay, M-monitor, Daegu, Korea) using eighty COVID-19 suspected clinical samples and compared these with the results of a commercial real-time reverse transcription polymerase chain reaction (RT-qPCR) assay (AllplexTM 2019-nCoV rRT-QPCR Assay, SeeGene, Seoul, Korea). The results of the RRT-LAMP assay targeting the N or RdRp gene of SARS-CoV-2 showed perfect agreement with the RT-qPCR assay results in terms of detection. Furthermore, the RRT-LAMP assay was completed in just within a 20-min reaction time, which is significantly faster than about the 2 h currently required for the RT-qPCR assay, thus enabling prompt decision making regarding the isolation of infected patients. The RRT-LAMP assay will be a valuable tool for rapid, sensitive, and specific detection of SARS-CoV-2 in human or unexpected animal clinical cases.

Effects of Curcuma longa L.on Human Stomach Cancer Cells (울금(鬱金)이 위암세포(胃癌細胞)에 미치는 영향(影響))

  • Cho, Yu-Kyung;Yoon, Song-Ryub;Kim, Beong-Woo;Kim, Jin-Sung;Ryu, Ki-Won;Ryu, Bong-Ha
    • THE JOURNAL OF KOREAN ORIENTAL ONCOLOGY
    • /
    • v.9 no.1
    • /
    • pp.15-37
    • /
    • 2003
  • Objective: We are aimed to identify anti-tumor effects of Curcuma longa L. on the stomach cancer cells through molecular biologic methods. Material & Methods: We used AGS as human stomach cancer cells obtained from American Type Culture Collection. The boiled extract of Curcuma longa L. $5{\mu}l$ (Sample I), $10{\mu}l$ (Sample II) was treated to cultural media(ml) for 0, 6, 12, 24, 48 hours. We measured the killing effect on stomach cancer cells through Trypan blue exclusion test and the suppressive effect on viability of stomach cancer cells via MTT assay. For identification of its anticancer mechanism, the revelation of Bcl-2, Bcl-XL, and Bax which are genes related to apoptosis using the quantitative RT-PCR, change of mitochondria membrane permeability and membrane potential via flow cytometry, the cycle of cell mitosis, caspase cleavage and annexin V staining were examined. Results: 1. showed significant killing effect on stomach cancer cell than the control group with a time(6 hours later) and density dependent manner, which was statistical significance. 2. Extract of Curcuma longa L. showed suppressive effect on viability of stomach cancer cells that each test groups had more suppressive effects on viability of stomach cancer cells than the control group with a time(6 hours later), which was statistical significance.(p<0.05) 3. In the test about the revelation of genes related to apoptosis, the revelation of Bcl-2 and Bcl-XL decreased with a density manner which was statistical significance. but the revelation of Bax was not changed with statistical significance. 4. Extract of Curcuma longa L. caused apoptosis by decreasing the absorbance of mitochondria with statistical significance, and also induced apoptosis by decreasing the membrane potential of mitochondria. 5. Extract of Curcuma longa L. destructed the cell cycle of cell mitosos. 6. Cell apoptosis was induced by extract of Curcuma longa L. certificated by method of caspase cleavage and annexin V staining. Conclusion: This experiment showed that Curcuma longa L. has anti-tumor effect with statistical significance. This is in vitro experiment and basic experiment on Curcuma longa L.. We hope more progressive research on Curcuma longa L. will go on and its anti-tumor effects will be more practically identified.

  • PDF