• 제목/요약/키워드: PCR condition

검색결과 338건 처리시간 0.03초

Association of a Newly Identified Variant of DNA Polymerase Beta (polβΔ63-123, 208-304) with the Risk Factor of Ovarian Carcinoma in India

  • Khanra, Kalyani;Bhattacharya, Chandan;Bhattacharyya, Nandan
    • Asian Pacific Journal of Cancer Prevention
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    • 제13권5호
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    • pp.1999-2002
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    • 2012
  • Background: DNA polymerase is a single-copy gene that is considered to be part of the DNA repair machinery in mammalian cells. The encoded enzyme is a key to the base excision repair (BER) pathway. It is evident that pol beta has mutations in various cancer samples, but little is known about ovarian cancer. Aim: Identification of any variant form of $pol{\beta}$ cDNA in ovarian carcinoma and determination of association between the polymorphism and ovarian cancer risk in Indian patients. We used 152 samples to isolate and perform RT-PCR and sequencing. Results: A variant of polymerase beta (deletion of exon 4-6 and 11-13, comprising of amino acid 63-123, and 208-304) is detected in heterozygous condition. The product size of this variant is 532 bp while wild type pol beta is 1 kb. Our study of association between the variant and the endometrioid type shows that it is a statistically significant factor for ovarian cancer [OR=31.9 (4.12-246.25) with p<0.001]. The association between variant and stage IV patients further indicated risk (${\chi}^2$ value of 29.7, and OR value 6.77 with 95% CI values 3.3-13.86). The correlation study also confirms the association data (Pearson correlation values for variant/stage IV and variant/endometrioid of 0.44 and 0.39). Conclusion: Individuals from this part of India with this type of variant may be at risk of stage IV, endometrioid type ovarian carcinoma.

Sphingobacterium sp. SW-09에 의한 토양환경에서의 다환 방향족탄화수소인 페난스렌의 분해 (Sphingobacterium sp. SW-09 Effectively Degrades Phenanthrene, a Polycyclic Aromatic Hydrocarbon, in a Soil Microcosm)

  • 손승우;장혜원;김성국;장종수
    • 생명과학회지
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    • 제21권11호
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    • pp.1511-1517
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    • 2011
  • 페난스렌은 다환방향족 탄화수소의 일종으로서 미량으로도 인체에 강한 해를 미칠 수 있는 주요 환경오염 물질이다. 미생물을 이용한 페난스렌 제거 목적으로 중국 쑤저우(Suzhou) 지역의 유류 오염토양에서 페난스렌을 강력하게 분해하는 세균을 분리하였다. 16S rDNA 염기서열 결정에 의하여 이 세균은 Sphingobacterium sp. SW-09로 동정되었으며 PCR 증폭을 통하여 페난스렌 분해 유전자인 nahH를 가지고 있음이 확인되었다. 이전의 연구에서 포천일대의 군부대에서 분리된 강력한 페난스렌 분해세균인 Staphylococcus sp. KW-07과 이번에 분리된 Sphingobacterium sp. SW-09을 이용하여 이들의 페난스렌 분해능을 비교분석하였다. 그 결과, 쑤저우 지역에서 분리된 Sphingobacterium sp. SW-09이 최소배지와 실제토양에서 모두 Staphylococcus sp. KW-07보다 강하게 페난스렌을 분해하는 것으로 나타났다. 결과적으로 이번에 분리된 Sphingobacterium sp. SW-09을 사용하여 유류 오염토양의 환경정화에 사용할 수 있을 것으로 판단된다.

염전으로부터 농화배양된 호염 메틸영양미생물 군집의 특성 (Prokaryotic Communities of Halophilic Methylotrophs Enriched from a Solar Saltern)

  • 김종걸;박수제;이성근
    • 미생물학회지
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    • 제46권3호
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    • pp.286-290
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    • 2010
  • C-1화합물은 고염분성 환경의 혐기적인 퇴적층에서 관찰되며, 이 퇴적층의 표면과 수면에는 호기성 메틸영양미생물의 잠재적인 서식지가 된다. 염전과 갯벌에서 채취한 토양 시료를 접종원으로 하여 메탄올을 탄소원과 에너지원으로 공급하고 염분농도에 따라 계대배양한 후 메탄올 산화 세균 성장 조건을 살펴 본 결과, 메탄올 산화 세균이 성장 할 수 있는 염분의 최대 농도는 20% 조건이었다. 변성 구배 젤 전기영동 (Denaturing gel gradient electrophoresis)을 이용하여 농화배양액 내 미생물 군집구조를 분석한 결과, 메탄올 산화 미생물인 Methylophaga 관련 세균이 우점하는 것으로 나타났다. 정량 PCR결과 고세균이 세균의 1-10%로 존재하는 것으로 나타났다. 세균용 항생제 실험결과, 메탄올 산화가 억제되어 고세균이 메탄올 산화에 관여하지 않는다는 것을 추정할 수 있었다. 이번 연구를 통해, 메틸영양세균이 고염분환경(염분 농도 20%까지)에서도 C-1 화합물을 산화할 수 있음을 확인 할 수 있었다.

북방전복 (Haliotis discus hannai) 의 대식세포이동저해인자 (MIF, Macrophage migration inhibitory factor) 유전자 동정 및 발현분석 (Cloning, characterization, and expression of the macrophage migration inhibitory factor gene from the pacific abalone (Haliotis discus hannai))

  • 박은희;신은하;김영옥;김동균;공희정;김우진;안철민;남보혜
    • 한국패류학회지
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    • 제32권4호
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    • pp.241-247
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    • 2016
  • Macrophage Migration Inhibitory Factor (MIF) are well-defined role as unique cytokine and critical mediator in acute and chronic inflammatory diseases, autoimmune diseases. In this study, we isolated and characterized a full-length of MIF cDNA from the abalone (Haliotis discus hannai). The full-length cDNA of abMIF was of 1264 bp, consisting of a 5'-terminal UTR of 143 bp, an open reading frame of 360 bp and a 3-terminal UTR of 761 bp. The abalone MIF cDNA encodes a 119-amino acid polypeptide with a calculated molecular mass of 13.4 kDa and isoelectric point of 9.07. Multiple alignments and phylogenetic analysis with the deduced abalone MIF protein and showed strong homology with disk abalone (Haliotis discusdiscus). The deduced amino acid sequence of abMIF exhibited homology with other reported MIFs, such as 80%, with that of other disk abalone H. discus discus MIF gene. Quantitative real-time PCR (qRT-PCR) analysis indicated that abMIF was highly expression observed in hapatopacreas, intestine, foot, and gonad of normal conditioned abalone. Even though AbMIF mRNA level in hemocytes was low under the normal condition, it was sharply up-regulated and reached the maximum at 6 h post-infection with Vibrio parahaemolyticus, and then decreased at 24 h post-infection. This result indicates that abMIF plays an important role in responding in the innate immune system.

Prevalence of Porphyromonas gingivalis fimA genotypes in peri-implant sulcus of Koreans using new primer

  • Kim, Sung-Geun;Hong, Ji-Youn;Shin, Seung-Il;Moon, Ji-Hoi;Lee, Jin-Yong;Herr, Yeek
    • Journal of Periodontal and Implant Science
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    • 제46권1호
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    • pp.35-45
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    • 2016
  • Purpose: Porphyromonas gingivalis fimA is a virulence factor associated with periodontal diseases, but its role in the pathogenesis of peri-implantitis remains unclear. We aimed to evaluate the relationship between the condition of peri-implant tissue and the distribution of P. gingivalis fimA genotypes in Koreans using a new primer. Methods: A total of 248 plaque samples were taken from the peri-implant sulci of 184 subjects. The control group consisted of sound implants with a peri-implant probing depth (PD) of 5 mm or less with no bleeding on probing (BOP). Test group I consisted of implants with a peri-implant PD of 5 mm or less and BOP, and test group II consisted of implants with a peri-implant PD of more than 5 mm and BOP. DNA was extracted from each sample and analyzed a using a polymerase chain reaction (PCR) with P. gingivalis -specific primers, followed by an additional PCR assay to differentiate the fimA genotypes in P. gingivalis-positive subjects. Results: The Prevalence of P. gingivalis in each group did not significantly differ (P>0.05). The most predominant fimA genotype in all groups was type II. The prevalence of type Ib fimA was significantly greater in test group II than in the control group (P<0.05). Conclusions: The fimA type Ib genotype of P. gingivalis was found to play a critical role in the destruction of peri-implant tissue, suggesting that it may be a distinct risk factor for periimplantitis.

형질전환된 상추내에서 GUS 유전자의 발현 및 후대검정 (Expression of $\beta$-Glucuronidase (GUS) Gene in Transgenic Lettuce (Lactuca sativa L.) and Its Progeny Analysis)

  • CHUNG, Jae Dong;KIM, Chang Kil;KIM, Kyung Min
    • 식물조직배양학회지
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    • 제25권4호
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    • pp.225-229
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    • 1998
  • 상추의 종자 무균발아후 4일된 자엽조직을 GUS 유전자가 도입된 A. tumefaciens LBA 4404와 2일간 공동배양한 다음 0.1mg/L NAA, 1.0mg/L 2ip, 50mg/L kanamycin, 500mg/L carbenicillin이 첨가된 MS 배지에 배양하여 식물체를 재분화시켰다. PCR 분석결과 GUS 유전자가 형질전환된 식물체의 게놈상에 삽입되어 있음을 확인하였다. 해부학적 GUS 활성을 분석하여 형질전환된 식물체의 줄기, 잎 그리고 뿌리에서 GUS 유전자의 발현을 확인하였다. 형질전환체로 확인된 식물체를 자가수정시켜 얻어진 종자의 GUS 활성을 분석하여 GUS 유전자가 발현되는 것을 확인하였다.

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Mouse Nerve Growth Factor Facilitates the Growth of Interspinal Schwannoma Cells by Activating NGF Receptors

  • Liu, Shu Yi;Liu, Sheng Ze;Li, Yu;Chen, Shi
    • Journal of Korean Neurosurgical Society
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    • 제62권6호
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    • pp.626-634
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    • 2019
  • Objective : Nerve growth factor (NGF) is a member of the neurotrophic factor family and plays a vital role in the physiological processes of organisms, especially in the nervous system. Many recent studies have reported that NGF is also involved in the regulation of tumourigenesis by either promoting or suppressing tumor growth, which depends on the location and type of tumor. However, little is known regarding the effect of NGF on interspinal schwannoma (IS). In the present study, we aimed to explored whether mouse nerve growth factor (mNGF), which is widely used in the clinic, can influence the growth of interspinal schwannoma cells (ISCs) isolated from IS in vitro. Methods : ISCs were isolated, cultured and identified by S-100 with immunofluorescence analysis. S-100-positive cells were divided into five groups, and separately cultured with various concentrations of mNGF (0 [phosphate buffered saline, PBS], 40, 80, 160, and 320 ng/mL) for 24 hours. Western blot and quantantive real time polymerase chain reaction (PCR) were applied to detect tyrosine kinase A (TrkA) receptor and p75 neurotrophin receptor ($p75^{NTR}$) in each group. Crystal violet staining was selected to assess the effect of mNGF (160 ng/mL) on ISCs growth. Results : ISCs growth was enhanced by mNGF in a dose-dependent manner. The result of crystal violet staining revealed that it was significantly strengthened the cells growth kinetics when cultured with 160 ng/mL mNGF compared to PBS group. Western blot and quantantive real time PCR discovered that TrkA receptor and mRNA expression were both up-regualated under the condition of mNGF, expecially in 160 ng/mL, while the exoression of $p75^{NTR}$ demonstrated no difference among groups. Conclusion : From these data, we conclude that exogenous mNGF can facilitate ISC growth by activating both TrkA receptor and $p75^{NTR}$. In addition, patients who are suffering from IS should not be administered mNGF in the clinic.

Epizootic Infection by Trypanosoma vivax in Cattle from the State of Minas Gerais, Brazil

  • Reis, Matheus de Oliveira;Souza, Fernanda Rezende;Albuquerque, Adriana Silva;Monteiro, Fernanda;Oliveira, Luan Francisco dos Santos;Raymundo, Djeison Lutier;Wouters, Flademir;Wouters, Angelica Terezinha Barth;Peconick, Ana Paula;Varaschin, Mary Suzan
    • Parasites, Hosts and Diseases
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    • 제57권2호
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    • pp.191-195
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    • 2019
  • Trypanosomiasis is caused by a pathogenic protozoan of the genus Trypanosoma, being Trypanosoma vivax the most important agent for cattle. The aim of the present study was to demonstrate the expansion of T. vivax infection in different mesoregions of Minas Gerais, Brazil, and describe the clinicopathological findings of trypanosomiasis in cattle. The diagnosis was based on visualization of the parasite in blood smears and DNA detection of T. vivax in the blood of live cows and tissues of necropsied animals by the polymerase chain reaction (PCR). Thirty suspected herds were tested, of which 11 were positive for T. vivax. The most frequent clinical signs were anemia, apathy, drop in milk production, weight loss, reproductive disorders, and nervous signs. Concomitant diseases, such as malignant edema, pneumonia and increased cases of mastitis were associated with T. vivax infection. Three cows were necropsied and the most significant findings were low body condition score, pale mucous and spleen with white pulp hyperplasia. The results demonstrated the expansion of T. vivax infection in Minas Gerais, that PCR-associated blood smears are promising for diagnosis, and that other diseases often occur concomitantly to T. vivax infection in regions with trypanosomiasis in cattle.

Wall shear stress on vascular smooth muscle cells exerts angiogenic effects on extracranial arteriovenous malformations

  • Ryu, Jeong Yeop;Park, Tae Hyun;Lee, Joon Seok;Oh, Eun Jung;Kim, Hyun Mi;Lee, Seok-Jong;Lee, Jongmin;Lee, Sang Yub;Huh, Seung;Kim, Ji Yoon;Im, Saewon;Chung, Ho Yun
    • Archives of Plastic Surgery
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    • 제49권1호
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    • pp.115-120
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    • 2022
  • Background In addition to vascular endothelial cells, vascular smooth muscle cells (VSMCs) are subject to continuous shear stress because of blood circulation. The angiogenic properties of VSMCs in extracranial arteriovenous malformations (AVMs) may exceed those of normal blood vessels if the body responds more sensitively to mechanical stimuli. This study was performed to investigate the hypothesis that rapid angiogenesis may be achieved by mechanical shear stress. Methods VSMCs were obtained from six patients who had AVMs and six normal controls. The target genes were set to angiopoietin-2 (AGP2), aquaporin-1 (AQP1), and transforming growth factor-beta receptor 1 (TGFBR1). Reverse-transcriptase polymerase chain reaction (RT-PCR) and real-time PCR were implemented to identify the expression levels for target genes. Immunofluorescence was also conducted. Results Under the shear stress condition, mean relative quantity values of AGP2, AQP1, and TGFBR1 in AVM tissues were 1.927±0.528, 1.291±0.031, and 2.284±1.461 when compared with neutral conditions. The expression levels of all three genes in AVMs were higher than those in normal tissue except for AQP1 under shear stress conditions. Immunofluorescence also revealed increased staining of shear stress-induced genes in the normal tissue and in AVM tissue. Conclusions Shear stress made the VSMCs of AVMs more sensitive. Although the pathogenesis of AVMs remains unclear, our study showed that biomechanical stimulation imposed by shear stress may aggravate angiogenesis in AVMs.

정량 PCR을 이용한 비위생 매립지의 특정 세균 및 효소 유전자와 수질인자와의 상관관계 평가 (Comparative Assessment of Specific Genes of Bacteria and Enzyme over Water Quality Parameters by Quantitative PCR in Uncontrolled Landfill)

  • 한지선;성은혜;박헌주;김창균
    • 대한환경공학회지
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    • 제29권8호
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    • pp.895-903
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    • 2007
  • 매립지를 직접 생태학적으로 모니터링하는 방법을 개발하고자, 매립지 내의 생화학적 반응에 관여하는 세균들과 효소의 양을 정량함과 동시에 지하수 수질인자와 상호 연관성을 조사하여 생태학적 인자와의 연계 이용 가능성을 평가하였다. 이를 위하여 4개의 매립 종료된 비위생 매립지(천안(C), 원주(W), 논산(N), 평택(P) 매립지)에서 계절별로 지하수 시료를 채취하였으며 동시에 16S rDNA 방법을 사용하여 미생물 다양성을 분석하였다. 이를 기반으로, 매립지에서 주로 발견되는 세균과 효소를 대표하는 유전자를 정량하기 위한 특이 프라이머 쌍을 제작하였으며 상관계수에 기초하여 수질인자와 유전자 지표 인자간의 정량적 관련성을 비교하였다. 그 결과 DSR(황환원 세균) gene과 BOD(생화학적 산소요구량)사이의 상관관계는 0.8 이상인데 반해 NSR(질산화 세균-Nitrospira sp.) gene과 질산성 질소는 0.9 이상이었다. 안정화지표(BOD/COD)와 MTOT(메탄 산화 세균), MCR(Methyl coenzyme M reductase), Dde(Dechloromonas denitrificans) gene들은 0.8 이상의 상관관계를 가졌으나 3가 철과 Fli(Ferribacterium limineticm) gene은 0.7로 낮았다. MTOT gene의 경우, BOD/COD과의 관련성이 100%에 가깝게 높았다. 또한, 혐기성 유전자들(nirS-아질산 환훤효소, MCR, Dde, DSR)과 DO 역시 0.8 이상으로 나타나 일반적인 매립지 혐기성 반응들이 DO에 크게 의존함을 보였다. 결론적으로 분자생물학적 조사와 수질인자가 높은 상호연관성이 있었으며 real-time PCR이 전통적인 모니터링 인자들과 동시에 상호 보완적으로 모니터링에 사용됨으로써 매립지안정화 및 주변 영향을 평가하는데 효율적으로 사용 될 수 있음을 알 수 있었다.