• Title/Summary/Keyword: LATE-PCR

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Study on the Characteristics of Feather Developing Pattern and Morphology in Early- and Late-Feathering Korean Native Chickens (한국재래닭에 있어 조우성과 만우성 깃털의 발생 양상 및 형태적 특성 고찰)

  • Bang, Min Hee;Cho, Eun Jung;Cho, Chang Yeon;Sohn, Sea Hwan
    • Korean Journal of Poultry Science
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    • v.45 no.3
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    • pp.155-165
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    • 2018
  • Chicken feathers could be classified into early-feathering (EF) and late-feathering (LF) depending on the development and patterns of the wing and tail feathers. Currently, feather-sexing is a widely used chick sexing method in the industry. This study was carried out to suggest the method of classifying of EF and LF chicks to establish auto-sexing Korean native chicken (KNC) strains. The development and morphology of wing feathers and tail feathers in 856 KNCs from hatching to 55-days old were analyzed to classify EF and LF chicks. We also performed PCR analysis using K-specific gene primers to confirm the agreement between the phenotypes and genotypes of EF and LF chickens. In the results, the EF chicks had long primaries and coverts, and there was a significant difference in length between primaries and coverts. The LF chicks had shorter primaries and coverts than the EF chicks, and showed little difference in the length between primaries and coverts. LF chicks could be classified into four groups: LF-Less, LF-Scant, LF-Equal and LF-Reverse according to their wing feather patterns. EF chicks had 1.5 times longer primaries than LF chicks until they were 15-days old, but the lengths were almost the same at 50-days old. The tail feathers of the EF chicks were apparent at 5-days old, but those of the LF chicks were short and indefinite at that time. When EF and LF chicks were classified by the length of primaries being more or less than 9 mm, the classification accuracies for EF and LF chicks were 96.2% and 85.4%, respectively, compared to the PCR results. In conclusion, juvenile EF and LF KNC showed distinct differences in feather development and morphology, and could be easily distinguished at one day-old.

Increased Expression of TGF-β1 in Correlation with Liver Fibrosis during Echinococcus granulosus Infection in Mice

  • Liu, Yumei;Abudounnasier, Gulizhaer;Zhang, Taochun;Liu, Xuelei;Wang, Qian;Yan, Yi;Ding, Jianbing;Wen, Hao;Yimiti, Delixiati;Ma, Xiumin
    • Parasites, Hosts and Diseases
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    • v.54 no.4
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    • pp.519-525
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    • 2016
  • To investigate the potential role of transforming growth factor (TGF)-${\beta}1$ in liver fibrosis during Echinococcus granulosus infection, 96 BALB/c mice were randomly divided into 2 groups, experimental group infected by intraperitoneal injection with a metacestode suspension and control group given sterile physiological saline. The liver and blood samples were collected at days 2, 8, 30, 90, 180, and 270 post infection (PI), and the expression of TGF-${\beta}1$ mRNA and protein was determined by real-time quantitative RT-PCR and ELISA, respectively. We also evaluated the pathological changes in the liver during the infection using hematoxylin and eosin (H-E) and Masson staining of the liver sections. Pathological analysis of H-E stained infected liver sections revealed liver cell edema, bile duct proliferation, and structural damages of the liver as evidenced by not clearly visible lobular architecture of the infected liver, degeneration of liver cell vacuoles, and infiltration of lymphocytes at late stages of infection. The liver tissue sections from control mice remained normal. Masson staining showed worsening of liver fibrosis at the end stages of the infection. The levels of TGF-${\beta}1$ did not show significant changes at the early stages of infection, but there were significant increases in the levels of TGF-${\beta}1$ at the middle and late stages of infection (P<0.05). RT-PCR results showed that, when compared with the control group, TGF-${\beta}1$ mRNA was low and comparable with that in control mice at the early stages of infection, and that it was significantly increased at day 30 PI and remained at high levels until day 270 PI (P<0.05). The results of this study suggested that increased expression of TGF-${\beta}1$ during E. granulosus infection may play a significant role in liver fibrosis associated with E. granulosus infection.

The Effects of Oviduct and Uterine Epithelial Cells on the Expression of Interleukin-$1\beta$ Gene in Preimplantation Mouse Embryos (생쥐 초기배아에서 Interleukin-$1\beta$ 유전자의 발현에 미치는 수란관과 자궁내막세포의 영향)

  • 홍석호;계명찬;김종월;이정복;오은정;조동제;최규완;김문규
    • Development and Reproduction
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    • v.3 no.1
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    • pp.59-67
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    • 1999
  • To investigate the role of interleukin-l$\beta$ (IL-1$\beta$) in the embryonic development, in vivo and in vitro expression patterns of IL-1$\beta$ gene in the preimplantation mouse embryos were examined by RT-PCR, and the effects of explanted mouse ovi-duct and uterine epithelial cells on the expression of IL-1$\beta$ gene in the pleimplantation mouse embryos were examined by co-culture. IL-1$\beta$ mRNA was detected in the embryos from 4-cell stage to blastocyst stage in vivo and from morula stage to hatching blastocyst stage in vitro. This transcript was not detected from the GV stage to late 2-cell stage in vivo, and not at the 4-cell and 8-cell stages in vitro. For the co-culture of late 2-cell embryos with the explanted mouse oviduct and uterine epithelial cells, oviducts and uterine epithelial cells were isolated at 48 hour alter the hCG injection. The explanted oviduct and uterine epithelial cells in co-culture groups facilitated the IL-1$\beta$ gene expression of the mouse embryos in comparison with the control. Taken together these results suggest that the presence of IL-1$\beta$ plays an important role in preimplantation embryonic development. In addition, the up-regulation of IL-1$\beta$ gene expression by the explanted oviduct and uterine epithelial cells demonstrates that embryonic expression of IL-l$\beta$ gene may be regulated by the interaction with oviductal and uterine factor (s).

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Calcium Mobilization Inhibits Lipid Accumulation During the Late Adipogenesis via Suppression of PPARγ and LXRα Signalings

  • Kim, Seung-Jin;Choi, Ho-Jung;Jung, Chung-Hwan;Park, Sung-Soo;Cho, Seung-Rye;Oh, Se-Jong;Kim, Eung-Seok
    • Food Science of Animal Resources
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    • v.30 no.5
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    • pp.787-794
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    • 2010
  • Calcium plays a role as a signaling molecule in various cellular events. It has been reported that calcium suppresses adipocyte differentiation only in the early phase of adipogenesis. Herein, we demonstrate that treatment of A23187, a mobilizer of intracellular calcium, on day 4 post adipocyte differentiation could still reduce lipid accumulation in differentiating 3T3-L1 cells for 48 h. In addition, luciferase reporter gene and RT-Q-PCR assays demonstrate that A23187 can selectively inhibit transcriptional activities and expression of PPAR$\gamma$ and LXR$\alpha$, suggesting that A23187 may reduce lipid accumulation in the late phase of adipogenesis via downregulation of PPAR$\gamma$ and LXR$\alpha$ expression and transactivation. Moreover, inhibition of HDAC activity by trichostatin A (TSA) partially blocked A23187-mediated downregulation of transcriptional activities of PPAR$\gamma$ and LXR$\alpha$. Together, our data demonstrate that calcium mobilization inhibits expression and transcriptional activities of PPAR$\gamma$ and LXR$\alpha$, resulting in reduced lipid accumulation in differentiating adipocytes, and thus, mobilization of intracellular calcium in adipocytes may serve as a new preventive and therapeutic approach for obesity.

The difference of castration timing of Korean Hanwoo bulls does not significantly affect the carcass characteristics

  • Hong, Heeok;Baatar, Delgerzul;Hwang, Seong-Gu
    • Journal of Animal Science and Technology
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    • v.63 no.2
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    • pp.426-439
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    • 2021
  • It is already well known that castration improves marbling quality but exact timing of castration is still highly debated in beef cattle production industry. After castration, blood hormonal changes occur in steer and objective of this study was to investigate the effects of growth hormone (GH) levels on adipocyte differentiation in stromal vascular cells (SVCs) and transdifferentiation into adipocytes in C2C12 myoblasts. Total GH concentrations were measured via enzyme-linked immunosorbent assay (ELISA) in 24 male calves and 4 female calves. Cell proliferation, cellular triglyceride (TG) accumulation, and the cell's lipolytic capability were measured in C2C12 myoblasts and SVCs. Myogenic, adipogenic, and brown adipocyte-specific gene expression was measured via real-time polymerase chain reaction (PCR) using SYBR green. Serum GH levels were the highest in late-castrated calves. Treatment with 5 ng/mL GH resulted in greater TG accumulation as well as increased CCAAT-enhancer-binding protein (C/EBP)α and peroxisome proliferator-activated receptor (PPAR)γ expression compared to that after treatment with 15 ng/mL GH. Treatment with 5 ng/mL GH also resulted in lower myogenin (myo)G and myoD expression compared to that after treatment with 15 ng/mL GH. The expression of bone morphogenetic protein (BMP) 7 after treatment with 5 ng/mL GH was higher than that after treatment with 15 ng/mL GH. But carcass characteristics data showed no significant difference between early and late castrated steers. Therefore, our results indicate that castration timing does not seem to be inevitable determinate of carcass qualities, particularly carcass weight and marbling score in Hanwoo beef cattle.

Characterization of CCND1 and TWIST1 as Prognostic Markers with the Mortality Rate of Breast Cancer

  • Ahn, Sungwoo;Park, Sangjung;Wang, Hye-Young;Park, Sunyoung;Kim, Jungho;Lee, Hyeyoung
    • Biomedical Science Letters
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    • v.24 no.2
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    • pp.76-86
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    • 2018
  • Breast cancer is one of the most common cancers affecting women worldwide. Although the survival rate of breast cancer has increased, breast cancer still results in a high mortality rate. Breast cancer deaths are caused by metastasis that occurs in organ dysfunction. Recently, there have been many studies on circulating tumor cells (CTCs), which are related to breast cancer metastasis in the blood. Recent studies have demonstrated that some CTCs do not express epithelial markers. Therefore, in this study, total RNA was extracted from blood without separating out the CTCs, and the characteristics of the CTCs were analyzed by RT-qPCR. Cyclin D1 and twist-related protein 1 (TWIST1) are well-known markers for predicting the prognosis of patients with breast cancer. However, few studies have demonstrated the use of CCND1 and TWIST1 in blood as diagnostic and prognostic markers of breast cancer. In this study, patients with late-stage breast cancer had overexpressed CCND1 and TWIST1 than patients with different stages of breast cancer (P < 0.001 and P < 0.01, respectively). The relative expression level of CCND1 in survivors was higher than in patients who died (P = 0.06). The relative expression level of TWIST1 in survivors was lower than in patients who died (P = 0.08). Overall CCND1 and TWIST1 were not useful as markers for the diagnosis of breast cancer through blood. However, we showed the possibility of using CCND1 and TWIST1 as prognostic markers, and a large-scale study is needed to confirm the usefulness of these prognostic markers.

Effect of Perillae Folium Extract on Regulation of Type 1 Allergic Response in RBL-2H3 Cells (자소엽(紫蘇葉) 에탄올 추출물이 RBL-2H3 비만세포에서 제 1형 알레르기 반응 조절에 미치는 효과)

  • Gok, Su-Yeong;Yu, Sun-Ae;Lee, Seung-Yeon
    • The Journal of Pediatrics of Korean Medicine
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    • v.26 no.1
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    • pp.36-45
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    • 2012
  • Objectives Perillae Folium (PF) has been widely used in Korean herbal medicines used for treatment of acute and chronic inflammatory diseases, such as rhinitis, asthma, and enteritis. In this study, to investigate the protective effect of PF on type 1 allergic response, we determined whether PF inhibits early or late allergic responses. Methods The effect of PF was analyzed by ELISA,. RT-PCR and Western blot in RBL-2H3 cells. Levels of ${\beta}$-hexosaminidase, interleukin (IL)-4 and TNF-${\alpha}$ were measured using enzyme-linked immunosorbent assays (ELISAs). mRNA levels of cytokines and enzymes were analyzed with RT-PCR. Signal transduction was analyzed with Western blot. Results We found that PF suppressed ${\beta}$-hexosaminidase release in RBL-2H3 by the IgE-DNP-HSA stimulation. PF also significantly inhibited enzymes level, such as COX-1, COX-2, iNOS, and HDC2, along with reduced cytokine levels, such as IL-2, IL-3, IL-4, IL-6, IL-13, and TNF-${\alpha}$ in RBL-2H3. In addition, PF suppressed the phospholyation of ERK1/2, JNK1/2, and $I{\kappa}B{\alpha}$. Conclusions Our results indicate that PF protects against type 1 allergic response and exert an anti-inflammatory effect through the inhibition of degranulation and expression of cytokines and enzymes via the suppression of signal transduction.

Role of Alkaline Serine Protease, Asp, in Vibrio alginolyticus Virulence and Regulation of Its Expression by LuxO-LuxR Regulatory System

  • Rui, Haopeng;Liu, Qin;Wang, Qiyao;Ma, Yue;Liu, Huan;Shi, Cunbin;Zhang, Yuanxing
    • Journal of Microbiology and Biotechnology
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    • v.19 no.5
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    • pp.431-438
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    • 2009
  • The alkaline serine protease asp, which was shown to be a virulence factor of Vibrio alginolyticus as a purified protein, was cloned from V. alginolyticus EPGS, a strain recently isolated from moribund Epinephelus coioides in an outbreak of vibriosis in a mariculture farm of Shenzhen. The asp null mutant was constructed by homologous recombination with suicide plasmid pNQ705-1. Compared with the wild-type strain, the asp null mutant exhibited a significant decrease of total extracellular protease activity, and caused a IS-fold decrease in virulence of V. alginolyticus. In our previous study, the luxO and $luxR_{val}$ genes from V. alginolyticus MVP01 were cloned and identified, and the luxO-$luxR_{val}$ regulatory couple was shown to regulate various genes expression, suggesting that it played a central role in the quorum sensing system of V. alginolyticus. In this study, the regulation of the asp gene was analyzed by using RT-PCR and quantitative real-time PCR methods; we proved that its transcription was greatly induced at the late stage of growth and was regulated by a luxO-$luxR_{val}$ regulatory system.

Inhibitory effect of Ulmus davidiana on ${\beta}$-hexosaminidase release and cytokine production in RBL-2H3 cells (유근피 추출물이 RBL-2H3 비만세포에서 ${\beta}$-hexosaminidase 및 cytokine 분비에 미치는 효과)

  • Park, Se-Bong;Kang, Kyung-Hwa;Yoon, Hwa-Jung;Ko, Woo-Shin
    • The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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    • v.24 no.1
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    • pp.86-95
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    • 2011
  • Objectives : Ulmus davidiana (UD) has been widely used in Korean herbal medicines used for treatment of acute and chronic inflammatory diseases, such as rhinitis, asthma, and abscess. In this study, To investigated the protective effect of UD on type 1 allergic response, we determined whether UD inhibits early and late allergic response. Methods : The effect of UD was analyzed by ELISA and RT-PCR in RBL-2H3 cells. Levels of ${\beta}$ -hexosaminidase, interleukin (IL)-4 and TNF-${\alpha}$ were measured using enzyme-linked immunosorbent assays (ELISAs). mRNA levels of COX-2 and T-helper type 2(Th2) cytokines were analyzed with RT-PCR. Results : We found that UD suppressed ${\beta}$-hexosaminidase release in RBL-2H3 not only by the PMA plus A23187 stimulation, but also by the IgE-DNP-HSA stimulation at the antigen-antibody binding stage and antibody-receptor binding stage. UD also significantly inhibited COX2 level, along with reduced Th2 cytokine levels, such as IL-3, IL-4, IL-5, IL-13, GM-CSF, and TNF-${\alpha}$ in RBL-2H3. Conclusions : Our results indicate that UD protects against type 1 allergic response and exerts an anti-inflammatory effect through the inhibition of degranulation and expression of COX2 and Th2 cytokines.

Spatio-Temporal Expression Pattern of Grp 78, a Putative Hoxc8 Downstream Target Gene, During Murine Embryogenesis

  • Kang Jin Joo;Kwon Yunjeong;Lee Eun Young;Park Hyoung Woo;Yang Hye-Won;Kim Myoung Hee
    • Biomedical Science Letters
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    • v.11 no.3
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    • pp.311-318
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    • 2005
  • Grp78, discovered as one of the putative target genes of Hoxc8, is a highly conserved stress protein and functions as a molecular chaperone in the endoplasmic reticulum (ER). In order to see the stage-specific expression pattern of Grp78 during development, mouse embryos from day 7.5 to 17.5 p.c. were isolated, and RT-PCR as well as in situ hybridization was performed. When RT-PCR was performed using Grp78 specific primers, periodic expression pattern was detected. And also a region-specific expression pattern was detected with a strong expression in the trunk part of day 11.5 p.c. embryo, like that of Hoxc8. When in situ hybridization was performed, Grp78 was revealed to be expressed in the endoderm, somite, neuroepithelium cells of neural tube in early embryos. In the case of late embryos, Grp78 expression was detected in the liver, segmental bronchus within cranial lobe of lung, ossification center within the cartilage primordium of rib and vertebra, submandibular gland, as well as metanephros. These expression patterns are very much similar to those of Hoxc8. Since Hoxc8 has been reported to regulate apoptosis during organogenesis, it might be possible that the apoptotic function could have been conveyed through the expression of Grp78, implying that the Grp78 is one of the Hoxc8 downstream target genes.

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