• 제목/요약/키워드: real-time TaqMan PCR

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수밀력 우수 꿀벌 계통 판별을 위한 계통 특이 분자마커 개발 (Identification of a Single Nucleotide Polymorphism (SNP) Marker for the Detection of Enhanced Honey Production in Hoenybee)

  • 김혜경;이명렬;이만영;최용수;김동원;강아랑
    • 한국양봉학회지
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    • 제32권3호
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    • pp.147-154
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    • 2017
  • 꿀벌은 화분매개 뿐만 아니라, 양봉산물을 생산하는 주요한 산업곤충 중 하나이다. 최근 농촌진흥청과 예천곤충 연구소에서는 국내 최초로 수밀력 우수 꿀벌 품종인 '장원벌'을 선발하여 보급하고 있다. 본 연구에서는 장원벌 계통 특이 분자 마커 개발을 위해 장원벌 부계인 D계통 특이적인 분자 마커를 개발 하고자 Sequence-Based Genotyping (SBG) 분석을 수행하였다. SGB 분석은 농촌진흥청 국립농업과학원에서 보존 육성중인 A, C, D, E, F 5개 기본종 계통에 대해 수행되었으며, 이를 통해 1,029개 SNP를 확보 할 수 있었다. 이후 A, C, D, F, E 기본종 계통 및 $D{\times}F$ 교배계통에 대한 SNP filtering 및 validation을 통해 최종적으로 AmD6 및 AmD9 두 개의 SNP 마커를 선발 하였으며, genotyping 분석을 통해 AmD9 마커가 장원벌 부계인 D 계통을 100% 구분 할 수 있는 것으로 확인되었다. 본 마커를 통해 D 계통 및 장원벌을 보다 정확하게 판별하고 육종에 활용 할 수 있을 것으로 기대하고 있다.

Predictive Role of ERCC1 and XPD Genetic Polymorphisms in Survival of Chinese Non-small Cell Lung Cancer Patients Receiving Chemotherapy

  • Zhang, Zhen-Yong;Tian, Xin;Wu, Rong;Liang, Yuan;Jin, Xue-Ying
    • Asian Pacific Journal of Cancer Prevention
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    • 제13권6호
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    • pp.2583-2586
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    • 2012
  • Aim: There is increasing evidence that ERCC1 and XPD have roles in response to chemotherapy among patients with NSCLC, but the results are conflicting. Therefore, we conducted the present prospective study in a Chinese population. Methods: A total of 632 primary NSCLC patients were included, followed-up from May 2006 to May 2011. Polymorphisms were detected by real time PCR with TaqMan probse, using genomic DNA extracted from peripheral blood samples. The Cox regression model was used to analyze the hazard ratios (HR) for ERCC1 and XPD. Results: The median time of follow-up was 31.6 months. Our results showed the ERCC1 118 T/T(HR=1.65, 95% CI=1.17-2.43) and XPD 751 Gln/Gln genotypes (HR=1.52, 95%CI=1.04-2.08) were associated with an increased risk of death from NSCLC. Moreover, the ERCC118 T allele and XPD 751 Gln allele genotypes had a more higher risk of death from NSCLC among both ex-smokers and current smokers. Conclusion: In summary, ERCC1 and XPD gene polymorphisms might provide better prognostic predictive information for NSCLC patients in Chinese populations, with smoking possibly interacting with the genotypes.

편도암의 발암 원인으로 Human Papilloma Virus를 통한 발암 기전과의 상관 관계 (Correaltion of Human Papilloma Virus Infection Status with Tonsillar Squamous Cell Carcinoma)

  • 김세헌;변형권;천제영;박영민;정진세;이소윤
    • 대한두경부종양학회지
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    • 제23권1호
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    • pp.21-25
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    • 2007
  • Background:Squamous cell carcinoma(SCC)of the palatine tonsils represents approximately 15-23% of all intraoral SCC. The most frequently reported risk factors for oropharyngeal cancer are smoking and alcohol. In a recent overview of HPV and tonsillar squamous cell carcinoma(TC), 51% contained HPV DNA, and HPV-16 being the most frequent type. We aimed to clarify whether HPV directly effects on the oncogenesis and biologic behavior of TC by comparison with infection prevalence, and physical status of virus. Material and Method:We used HPV genotyping DNA chip(Biocore, Korea, Seoul) arrayed by multiple oligonucleotide probes of L1 sequence of 26 types of HPV and HPV genotypes are identified by fluorescence scanner. The copy numbers of HPV E2 and E6 open reading frames(ORF) were assessed using a TaqMan-based 5'-exonuclease quantitative real-time PCR assay. The ratio of E2 to E6 copy numbers was calculated to determine the physical status of HPV-16 viral gene. Results:We observed a significant difference in HPV prevalence between 52 TCs and 69 CFTs(73.1% vs. 11.6%), and most of the HPVs were type 16(87.2%)and non-episomal(94.1%) state. Conclusions:This study regarding HPV infection prevalence and mechanism in the largest population of palatine tonsillar squamous cell carcinoma with chronic follicular tonsillitis revealed significant difference pf HPV prevalence between TC and CFT. Most of HPV were 16 type and integrated or mixed, HPV-16 integration could be directly related to tonsillar carcinogenesis.

Genetic Variations in XRCC4 (rs1805377) and ATF6 (rs2070150) are not Associated with Hepatocellular Carcinoma in Thai Patients with Hepatitis B Virus Infection

  • Makkoch, Jarika;Praianantathavorn, Kesmanee;Sopipong, Watanyoo;Chuaypen, Natthaya;Tangkijvanich, Pisit;Payungporn, Sunchai
    • Asian Pacific Journal of Cancer Prevention
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    • 제17권2호
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    • pp.591-595
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    • 2016
  • The liver is one of the most common sites of cancer in the world, hepatocellular carcinoma (HCC) predominating. Chronic hepatitis B virus infection (CHB) is considered as an important potential risk factors for HCC. Different people have diverse responses to HBV infection regarding the likelihood of HCC development, and host factors such as single nucleotide polymorphisms (SNPs) might account for this. The present study was conducted to evaluate any association between SNP frequencies in two genes, XRCC4 (rs1805377) and ATF6 (rs2070150), and the risk of CHB and HCC development in Thai patients. The study covered 369 subjects including 121 HCC patients, 141 with chronic hepatitis B virus infection (CHB) and 107 healthy controls. With TaqMan real-time PCR, the results showed that no significant association between XRCC4 (rs1805377) and ATF6 (rs2070150) and risk of HCC in the Thai population. From this first study of the 2 polymorphisms and HCC in Thailand it can concluded that rs1805377 and rs2070150 polymorphisms may not be applicable as genetic markers in the Thai population for HCC assessment.

Association of PNPLA3 Polymorphism with Hepatocellular Carcinoma Development and Prognosis in Viral and Non-Viral Chronic Liver Diseases

  • Khlaiphuengsin, Apichaya;Kiatbumrung, Rattanaporn;Payungporn, Sunchai;Pinjaroen, Nutcha;Tangkijvanich, Pisit
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권18호
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    • pp.8377-8382
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    • 2016
  • Background: The aim of this study was to evaluate any association between a single nucleotide polymorphism (SNP) in the patatin-like phospholipase domain containing 3 (PNPLA3) (rs738409, C>G) and the development and prognosis in patients with hepatocellular carcinoma (HCC). Materials and Methods: Two hundred heathy controls and 388 HCC cases were included: 211 with HBV, 98 patients with HCV, 29 with alcoholic steatohepatitis (ASH) and 52 with non-alcoholic steatohepatitis (NASH). The SNP was determined by real-time PCR based on TaqMan assays. Results: The prevalence of rs738409 genotypes CC, CG and GG in controls was 91 (45.5%), 88 (44.0%), and 21 (10.5%), respectively, while the corresponding genotypes in all patients with HCC was 158 (40.7%), 178 (45.9%), and 52 (13.4%). The GG genotype had significantly higher distribution in patients with ASH/NASH-related HCC compared with controls (OR=2.34, 95% CI=1.16-4.71, P=0.018), and viral-related HCC cases (OR=2.15, 95% CI=1.13-4.08, P=0.020). However, the frequency of the GG genotype was similar between controls and patients with viral-related HCC. At initial diagnosis, HBV-related HCC were larger and at more advanced BCLC stage than the other HCC groups. There were no significant differences between the GG and non-GG groups regarding clinical characteristics, tumor stage and overall survival. Conclusions: These data suggest an influence of the PNPLA3 polymorphism on the occurrence of HCC in patients with ASH/NASH but not among those with chronic viral hepatitis. However, the polymorphism was not associated with the prognosis of HCC.

Single Nucleotide Polymorphisms in miR-149 (rs2292832) and miR-101-1 (rs7536540) Are Not Associated with Hepatocellular Carcinoma in Thai Patients with Hepatitis B Virus Infection

  • Pratedrat, Pornpitra;Sopipong, Watanyoo;Makkoch, Jarika;Praianantathavorn, Kesmanee;Chuaypen, Natthaya;Tangkijvanich, Pisit;Payungporn, Sunchai
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권15호
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    • pp.6457-6461
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    • 2015
  • MicroRNAs directly and indirectly influence many biological processes such as apoptosis, cell maintenance, and immune responses, impacting on tumor genesis and metastasis. They modulate gene expression at the posttranscriptional level and are associated with progression of liver disease. Hepatocellular carcinoma (HCC) is a cancer which mostly occurs in males. There are many factors affect HCC development, for example, hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus (HIV), co-infection, environmental factors including alcohol, aflatoxin consumption and host-related factors such as age, gender immune response, microRNA and single nucleotide polymorphisms (SNPs). Chronic infection with the hepatitis B virus is the major factor leading to HCC progression since it causes the liver injury. At present, there are many reports regarding the association of SNPs on miRNAs and the HCC progression. In this research, we investigated the role of miR-149 (rs2292832) and miR-101-1 (rs7536540) with HCC progression in Thai population. The study included 289 Thai subjects including 104 HCC patients, 90 patients with chronic hepatitis B virus infection (CHB) and 95 healthy control subjects. The allele and genotype of rs2292832 and rs7536540 polymorphisms were determined by TaqMan real-time PCR assay. Our results revealed no significant association between miR-149 (rs2292832) and miR-101-1 (rs7536540) and the risk of HCC in our Thai population. However, this research is the first study of miR-149 (rs2292832) and miR-101-1 (rs7536540) in HCC in Thai populations and the results need to be confirmed with a larger population.

Single Nucleotide Polymorphism of Interferon Lambda-4 Gene is not Associated with Treatment Response to Pegylated Interferon in Thai Patients with Chronic Hepatitis B

  • Limothai, Umaporn;Wasitthankasem, Rujipat;Poovorawan, Yong;Tangkijvanich, Pisit
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권13호
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    • pp.5515-5519
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    • 2015
  • The single nucleotide polymorphism (SNP) ss469415590 in the interferon lambda-4 (IFNL4) gene has recently been reported to have an association with treatment response in chronic hepatitis C. However, any importance of the SNP in association with response to pegylated interferon (PEG-IFN) therapy in patients with chronic hepatitis B (CHB) is unclear. We retrospectively analyzed data for Thai patients with CHB treated with PEG-IFN for 48 weeks. Virological response (VR) for HBeAg-positive CHB was defined as HBeAg seroconversion plus HBV DNA level <2,000 IU/mL at 24 weeks post-treatment. VR for HBeAg-negative CHB was defined as an HBV DNA level <2,000 IU/mL at 48 weeks. The SNP was identified by real time PCR using the TaqMan genotyping assay with MGB probes. A total 254 patients (107 HBeAg-positive and 147 HBeAg-negative) were enrolled in the study. The distribution of TT/TT, ${\Delta}G/TT$ and ${\Delta}G/{\Delta}G$ genotypes was 221 (87.0%), 32 (12.6%) and 1 (0.4%), respectively. Patients with non-TT/TT genotypes had significantly higher baseline HBV DNA levels than patients with the TT/TT genotype. In HBeAg-positive CHB, 41.2% of patients with TT/TT genotype versus 50.0% with non-TT/TT genotype achieved VR (P=0.593). In HBeAg-negative CHB, the corresponding figures were 40.3% and 43.5%, respectively (P=0.777). There was no significant correlation between the SNP genotypes and HBsAg clearance in both groups of patients. In summary, ss469415590 genotypes were not associated with response to PEG-IFN in Thai patients with HBeAg-positive and HBeAg-negative CHB.

miR-200a Overexpression in Advanced Ovarian Carcinomas as a Prognostic Indicator

  • Zhu, Cheng-Liang;Gao, Guo-Sheng
    • Asian Pacific Journal of Cancer Prevention
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    • 제15권20호
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    • pp.8595-8601
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    • 2014
  • Background: miR-200a expression is frequently altered in numerous cancers. The aim of the present study was to determine the role of microRNA-200a in advanced ovarian carcinomas. Materials and Methods: We measured miR-200a expression in 72 matched normal ovarian tissues and advanced ovarian carcinomas, and also two ovarian carcinoma cell lines (SKOV3 and SKOV3.ip1 - the latter being more invasive and metastatic than the parental SKOV3) by stem-loop real-time RT-PCR based on TaqMan microRNA assay using U6 as a reference. Levels of miR-200a expression were compared by disease stage, tumor grade, histology, and lymph node involvement. To evaluate the role of microRNA-200a, cell proliferation and invasion of SKOV-3 and SKOV-3.ip1 were analyzed with miR-200a inhibitor/mimic transfected cells. Results: Of 72 paired samples, 65 cancer tissues overexpressed microRNA-200a greater than two fold in comparison with matched normal epithelium. Specifically, patients with lymph node metastasis showed significant elevation. The level correlated with clinicopathological features, including high tumor grade, late disease stage, most notably with lymph node metastasis, but not with tumor histology. In addition, SKOV-3.ip1 cells also overexpressed miR-200a compared with SKOV-3, and miR-200a inhibitor transfected SKOV-3.ip1 cells showed significant reduction in cellular proliferation and invasion, while a miR-200a mimic stimulated the opposite behavior. Conclusions: We provide definitive evidence that miR-200a is up-regulated in a significant proportion of advanced ovarian carcinomas, and that elevated miR-200a expression facilitates tumor progression. Our findings support the notion that miR-200a is an onco-microRNA for ovarian cancer, and elevation is a useful potential diagnostic indicator. This study also provides a solid basis for further functional analysis of miR-200a in advanced ovarian cancer.

두경부 편평세포암종의 발암 원인으로 인간유두종 바이러스(Human Papilloma Virus)의 역할 및 이와 관련된 발암 기전에 관한 연구 (The Etiologic Roles and Carcinogenic Mechanisms of Human Papilloma Virus in Head and Neck Squamous Cell Carcinoma)

  • 신동현;이세영;구본석;김세헌
    • 대한두경부종양학회지
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    • 제25권1호
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    • pp.28-32
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    • 2009
  • Background : The most frequently reported risk factors for head and neck suamous cell carcinoma are smoking and alcohol. But in a recent overview, human papilloma virus(HPV) infection was revealed the important carcinogenic factor in oropharyngeal cancer. We aimed to clarify whether HPV directly effects on the oncogenesis and biologic behavior of hean and neck squamous cell carcinoma by comparison with infection prevalence, and physical status of virus. Material and Method : We used HPV genotyping DNA chip(Biocore, Korea, Seoul) arrayed by multiple oligonucleotide probes of L1 sequence of 26 types of HPV and HPV genotypes are identified by fluorescence scanner. The copy numbers of HPV E2 and E6 open reading frames(ORF) were assessed using a TaqMan-based 5'-exonuclease quantitative real-time PCR assay. The ratio of E2 to E6 copy numbers was calculated to determine the physical status of HPV-16 viral gene. Results : We observed a significant difference in HPV prevalence between tonsillar cancer group and control group(73.1% vs. 11.6%), and most of the HPVs were type 16(87.2%) and integrated(94.1%) state. In terms of oral tongue cancer, we demonstrate that 30.5% has integrated HPV-16 in cancer tissue. But Glottic cancer only 1% is related to HPV-16 integration. Conclusion : This study revealed significant relationship of HPV prevalence with oropharyngeal and oral tongue squamous cell carcinoma. Most of HPV were 16 type and integrated or mixed, HPV-16 integration could be directly related to the carcinogenesis.

만성 피부 염증소견을 보인 개의 면역학적 특성 분석 연구 (Analysis of Immune Response in Dogs with Chronic Inflammatory Skin Disease)

  • 조선주;고민수;정복기;고재형;윤소라;한동운;이봉주
    • 한국임상수의학회지
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    • 제26권5호
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    • pp.433-440
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    • 2009
  • High levels of inflammatory cytokines were proposed contributors to the pathogenesis of a various inflammatory skin disorders. Therefore, investigating the immune response of the inflammatory skin disorder allows a better understanding of pathogenesis of a various inflammatory skin disorders and therapeutic approaches. The aim of this study was to analyze of the immune response in dogs with chronic inflammatory skin disease. To this aim, the present study evaluated relative mRNA expression of canine $IFN-{\gamma}$, IL-4, $TGF-{\beta}$ and IL-10 using TaqMan realtime PCR assays and semi-quantitative RT-PCR in freshly isolated peripheral blood mononuclear cells from twenty dogs with chronic inflammatory skin disease and ten normal dogs. The relative mRNA expression levels of IL-4 mRNA were significantly higher in dogs with chronic inflammatory skin disease than those in normal dogs (P < 0.01). The results of present study also showed a tendency towards increased expression of IL-10 transcripts in dogs with chronic inflammatory skin disease. However, there were no significant differences in the levels $IFN-{\gamma},\;TGF-{\beta}$ between normal and chronically inflammed dogs. In addition, the concentration of serum IgE was significantly increased in dogs with chronic inflammatory skin disease compared with those in normal dogs (P < 0.01). In histopathological examination, we found that there were markedly increased mast cell counts in chronically inflammed dogs (P < 0.05). These results suggest that the pathogenesis of chronic inflammatory skin disease might be associated with a T-cell mediated inflammatory responses characterized by a Th2-skewed immune response. Based on these results, the modulation of Th1/Th2 balance may be an effective therapeutic strategy for the treatment of chronic inflammatory skin disease.