• Title/Summary/Keyword: Single PCR

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Vascular Endothelial Growth Factor (VEGF) Gene Polymorphisms and Breast Cancer Risk in a Chinese Population

  • Luo, Ting;Chen, Long;He, Ping;Hu, Qian-Cheng;Zhong, Xiao-Rong;Sun, Yu;Yang, Yuan-Fu;Tian, Ting-Lun;Zheng, Hong
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.4
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    • pp.2433-2437
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    • 2013
  • Vascular endothelial growth factor (VEGF) is a potent regulator of angiogenesis and thereby involved in the development and progression of solid tumours. Associations between three VEGF gene polymorphisms (-634 G/C, +936 C/T, and +1612 G/A) and breast cancer risk have been extensively studied, but the currently available results are inconclusive. Our aim was to investigate associations between three VEGF gene polymorphisms and breast cancer risk in Chinese Han patients. We performed a hospital-based case-control study including 680 female incident breast cancer patients and 680 female age-matched healthy control subjects. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis was performed to detect the three VEGF gene polymorphisms. We observed that women carriers of +936 TT genotypes [odds ratio (OR) =0.46, 95% confidence interval (CI) = 0.28, 0.76; P=0.002] or 936 T-allele (OR=0.81, 95% CI= 0.68, 0.98; P=0.03) had a protective effect concerning the disease. Our study suggested that the +1612G/A polymorphism was unlikely to be associated with breast cancer risk. The -634CC genotype was significantly associated with high tumor aggressiveness [large tumor size (OR=2.63, 95% CI=1.15, 6.02; P=0.02) and high histologic grade (OR=1.47, 95% CI= 1.06, 2.03; P=0.02)]. The genotypes were not related with other tumor characteristics such as regional or distant metastasis, stage at diagnosis, or estrogen or progesterone receptor status. Our study revealed that the VEGF -634 G/C and +936 C/T gene polymorphisms may be associated with breast cancer in Chinese Han patients.

Screening and Characterization of LTR Retrotransposons in the genomic DNA of Pleurotus eryngii (큰느타리버섯 유전체내 LTR Retrotransposon 유전자 탐색 및 특성연구)

  • Kim, Sinil;Le, Quy Vang;Kim, Sun-Mi;Ro, Hyeon-Su
    • The Korean Journal of Mycology
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    • v.42 no.1
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    • pp.50-56
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    • 2014
  • Transposable elements (TEs) are mobile DNA elements that often cause mutations in genes and alterations in the chromosome structure. In order to identify and characterize transposable elements (TEs) in Pleurotus eryngii, a TE-enriched library was constructed using two sets of TE-specific degenerated primers, which target conserved sequences of RT and RVE domains in fungal LTR retrotransposons. A total of 256 clones were randomly chosen from the library and their insert sequences were determined. Comparative investigation of the insert sequences with those in repeat element database, Repbase, revealed that 71 of them were found to be TE-related fragments with significant similarity to LTR retrotransposons from other species. Among the TE sequences, the 70 TEs were Gypsy-type LTR retrotransposons, including 20 of MarY1 from Tricholoma matsutake, 26 of Gypsy-8_SLL from Serpula lacrymans, and 16 of RMER17D_MM from mouse, whereas a single sequence, Copia-48-PTR, was found as only Copia-type LTR retrotransposon. Southern blot analysis of the HindIII-digested P. eryngii genomic DNA showed that the retrotransposon sequences similar to MarY1 and Gypsy-8_SLL were contained as high as 14 and 18 copies per genome, respectively, whereas other retrotransposons were remained low. Moreover, both of the two Gypsy retrotransposons were expressed in full length mRNA as shown by Northern blot analysis, suggesting that they were functionally active retrotransposons.

Molecular Genetic Identification of Yeast Strains Isolated from Egyptian Soils for Solubilization of Inorganic Phosphates and Growth Promotion of Corn Plants

  • Hesham, Abd El-Latif;Mohamed, Hashem M.
    • Journal of Microbiology and Biotechnology
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    • v.21 no.1
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    • pp.55-61
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    • 2011
  • Forty yeast strains isolated from soils taken from different locations in Egypt were tested for their P-solubilizing activities on the basis of analyzing the clear zone around colonies growing on a tricalcium phosphate medium after incubation for 5 days at $25^{\circ}C$, denoted as the solubilization index (SI). Nine isolates that exhibited P-solubilization potential with an SI ranging from 1.19 to 2.76 were genetically characterized as five yeasts belonging to the genus Saccharomyces cerevisiae and four non-Saccharomyces, based on a PCR analysis of the ITS1-26S region amplied by SC1/SC2 species-specific primers. The highest P-solubilization efficiency was demonstrated by isolate PSY- 4, which was identified as Saccharomyces cerevisiae by a sequence analysis of the variable D1/D2 domain of the 26S rDNA. The effects of single and mixed inoculations with yeast PSY-4 and Bacillus polymyxa on the P-uptake and growth of corn were tested in a greenhouse experiment using different levels of a phosphorus chemical fertilizer (50, 100, and 200 kg/ha super phosphate 15.5% $P_2O_5$). The results showed that inoculating the corn with yeast PSY-4 or B. polymyxa caused significant increases in the shoot and root dry weights and P-uptake in the shoots and roots. The P-fertilization level also had a significant influence on the shoot and root dry weights and P-uptake in the shoots and roots when increasing the P-level from 50 up to 200 kg/ha. Dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 200 kg/ha gave higher values for the shoot and root dry weights and P-uptake in the shoots and roots, yet these increases were nonsignificant when compared with dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha. The best increases were obtained from dual inoculation with yeast strain PSY-4 and B. polymyxa at a P-fertilization level of 100 kg/ha, which induced the following percentage increases in the shoot and root dry weights, and P-uptake in the shoots and roots; 16.22%, 46.92%, 10.09%, and 31.07%, respectively, when compared with the uninoculated control (fertilized with 100 kg/ha).

Lack of Influence of the SMAD7 Gene rs2337107 Polymorphism on Risk of Colorectal Cancer in an Iranian Population

  • Akbari, Zahra;Safari-Alighiarloo, Nahid;Haghighi, Mahdi Montazer;Vahedi, Mohsen;Mirtalebi, Hanieh;Azimzadeh, Pedram;Milanizadeh, Saman;Shemirani, Atena Irani;Nazemalhosseini-Mojarad, Ehsan;Aghdaei, Hamid Asadzadeh;Zali, Mohammad Reza
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.11
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    • pp.4437-4441
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    • 2014
  • SMAD7 has been identified as a functional candidate gene for colorectal cancer (CRC). SMAD7 protein is a known antagonist of the transforming growth factor beta ($TGF-{\beta}$) signaling pathway which is involved in tumorigenesis. Polymorphisms in SMAD7 may thus alter cancer risk. The aim of this study was to investigate the influence of a SMAD7 gene polymorphism (rs2337107) on risk of CRC and clinicopathological features in an Iranian population. In total, 210 subjects including 105 patients with colorectal cancer and 105 healthy controls were recruited in our study. All samples were genotyped by TaqMan assay via an ABI 7500 Real Time PCR System (Applied Biosystems) with DNA from peripheral blood. The polymorphism was statistically analyzed to investigate the relationship with the risk of colorectal cancer and clinicopathological properties. Logistic regression analysis revealed that there was no significant association between rs2337107and the risk of colorectal cancer. In addition, no significant association between genotypes and clinicopathological features was observed (p value>0.05). Although there was not any association between genotypes and disorder, CT was the most common genotype in this population. This genotype prevalence was also higher in the patients with well grade (54.9%) and colon (72.0%) tumors. Our results provide the first evidence that this polymorphism is not a potential contributor to the risk of colorectal cancer and clinicopathological features in an Iranian population, and suggests the need of a large-scale case-control study to validate our results.

Cytochrome P450 1A1, 2E1 and GSTM1 Gene Polymorphisms and Susceptibility to Colorectal Cancer in the Saudi Population

  • Saeed, Hesham Mahmoud;Alanazi, Mohammad Saud;Nounou, Howaida Attia;Shalaby, Manal Ali;Semlali, Abdelhabib;Azzam, Nahla;Aljebreen, Abdeulrahan;Alharby, Othman;Parine, Narasimha Reddy;Shaik, Jilani;Maha, Maha
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.6
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    • pp.3761-3768
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    • 2013
  • Background: The Saudi population has experienced a sharp increase in colorectal and gastric cancer incidences within the last few years. The relationship between gene polymorphisms of xenobiotic metabolizing enzymes and colorectal cancer (CRC) incidence has not previously investigated among the Saudi population. The aim of the present study was to investigate contributions of CYP1A1, CYP2E1, and GSTM1 gene polymorphisms. Materials and Methods: Blood samples were collected from CRC patients and healthy controls and genotypes were determined by polymerase chain reaction restriction fragment length polymorphism and sequencing. Results and Conclusions: $CYP2E1^*6$ was not significantly associated with CRC development (odd ratio=1.29; confidence interval 0.68-2.45). A remarkable and statistically significant association was observed among patients with $CYP1Awt/^*2A$ (odd ratio=3.65; 95% confidence interval 1.39-9.57). The $GSTM1^*0/^*0$ genotype was found in 2% of CRC patients under investigation. The levels of CYP1A1, CYP2E1 and GSTM1 mRNA gene expression were found to be 4, 4.2 and 4.8 fold, respectively, by quantitative real time PCR. The results of the present case-control study show that the studied Saudi population resembles Caucasians with respect to the considered polymorphisms. Investigation of genetic risk factors and susceptibility gene polymorphisms in our Saudi population should be helpful for better understanding of CRC etiology.

IL-6 gene promoter polymorph isms in Korean generalized aggressive periodontitis (한국인 전반적 급진성 치주염 환자에서 IL-6 유전자 다변성에 관한 연구)

  • Bang, Sun-Jung;Kim, Il-Shin;Kim, Ok-Su;Kim, Young-Jun;Chung, Hyun-Ju
    • Journal of Periodontal and Implant Science
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    • v.38 no.4
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    • pp.579-588
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    • 2008
  • Purpose: The purpose of this study was to investigate the association of generalized aggressive periodontitis with IL-6 promoter gene single nucleotide polymorphisms(SNP). Material and Methods: The study population consisted of 52 generalized aggressive periodontitis patients(GAP) and 30 periodontally healthy control subjects, who were systemically healthy non-smokers. Genomic DNA was obtained from buccal swab. The IL-6 promotor SNP at the positions of -597, -572, and -174 were genotyped by amplifying the polymorphic region using polymerase chain reaction(PCR), restriction enzyme digestion and gel electrophoresis. Result: The genotype distributions for G/G, G/A and A/A genotypes of IL-6 -597 were 30.8%, 40.4%, and 28.8% in the GAP group and 53.3%, 40%, and 6.7% in the control group and were statistically different between 2 groups(p<0.05). Allele 2 frequency of IL-6 -597 were significantly higher in the GAP group than the control group(p<0.01). At the position of IL-6 -572, the distribution for C/C, C/G and G/G genotypes were 23.1%, 55.8% and 21.2% in the GAP group and 20%, 33.3%, and 46.7% in the control group. In female subjects, the genotype distribution were significantly different between 2 groups(p<0.01). In male subjects, allele 2 frequency of IL-6-572 was significantly lower in the GAP group than the control group(p<0.05). The genotype distribution of IL-6 -174 in the GAP group were 96.2%, 3.8% for G/G, G/C genotypes whereas only the G/G genotype was detected in the control group. Conclusion: In conclusion, significant associations were found in IL-6 gene promoter(-597, -572) polymorphisms and generalized aggressive periodontitis. Further cohort study will be necessary in larger population.

Screening of Natural Product Libraries for the Extension of Cell Life-span through Immune System (면역시스템을 통한 세포수명연장 천연물질 스크린)

  • Yoo, Bo-Kyung;Kwon, Kisang;Ko, Young Hwa;Kim, Hong Geun;Lee, Seokhyun;Park, Kwan-Ho;Choi, Ji-Young;Kwon, O-Yu
    • Journal of Life Science
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    • v.26 no.3
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    • pp.359-363
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    • 2016
  • We have screened four natural products against 640 single compounds, which shows more two folds gene expression for both endoplasmic reticulum aminopeptidase 1 (ERAP1) and FOXO-family transcription factor (FOXO1). The results were as follows. (±)-Car-3-ene-2,5-dione from Asarum sieboldii Miq. is C10H12O2 molecular formula and the 164 kDa molecular weight. Cinobufagin from Bufonis Venennum is C26H34O6 molecular formula and 442 kDa molecular weight. So far reported main biological function is Na+/K+-ATPase inhibition. Corilagin from Euphorbia pekinensis is C27H22O18 molecular formula and 634 kDa molecular weight. Carbonic anhydrase inhibition is well known its biological function. Corydaline from Corydalis turtschaninovii is C22H27NO4 molecular formula and 369 kDa molecular weight. The main biological function is acetylcholinesterase inhibition. In the short future, four types of natural products will be used in longevity experiments with insects. The results may give one of the clues for studying new drug development candidates of the longevity.

Enhanced Expression of TREK-1 Is Related with Chronic Constriction Injury of Neuropathic Pain Mouse Model in Dorsal Root Ganglion

  • Han, Hyo Jo;Lee, Seung Wook;Kim, Gyu-Tae;Kim, Eun-Jin;Kwon, Byeonghun;Kang, Dawon;Kim, Hyun Jeong;Seo, Kwang-Suk
    • Biomolecules & Therapeutics
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    • v.24 no.3
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    • pp.252-259
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    • 2016
  • Neuropathic pain is a complex state showing increased pain response with dysfunctional inhibitory neurotransmission. The TREK family, one of the two pore domain $K^+$ (K2P) channel subgroups were focused among various mechanisms of neuropathic pain. These channels influence neuronal excitability and are thought to be related in mechano/thermosensation. However, only a little is known about the expression and role of TREK-1 and TREK-2, in neuropathic pain. It is performed to know whether TREK-1 and/or 2 are positively related in dorsal root ganglion (DRG) of a mouse neuropathic pain model, the chronic constriction injury (CCI) model. Following this purpose, Reverse Transcription Polymerase Chain Reaction (RT-PCR) and western blot analyses were performed using mouse DRG of CCI model and compared to the sham surgery group. Immunofluorescence staining of isolectin-B4 (IB4) and TREK were performed. Electrophysiological recordings of single channel currents were analyzed to obtain the information about the channel. Interactions with known TREK activators were tested to confirm the expression. While both TREK-1 and TREK-2 mRNA were significantly overexpressed in DRG of CCI mice, only TREK-1 showed significant increase (~9 fold) in western blot analysis. The TREK-1-like channel recorded in DRG neurons of the CCI mouse showed similar current-voltage relationship and conductance to TREK-1. It was easily activated by low pH solution (pH 6.3), negative pressure, and riluzole. Immunofluorescence images showed the expression of TREK-1 was stronger compared to TREK-2 on IB4 positive neurons. These results suggest that modulation of the TREK-1 channel may have beneficial analgesic effects in neuropathic pain patients.

Genetic Differences within and between Populations of Korean Catfish (S. asotus) and Bullhead (P. fulvidraco) Analysed by RAPD-PCR

  • Yoon, Jong-Man;Kim, Jong-Yeon
    • Asian-Australasian Journal of Animal Sciences
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    • v.17 no.8
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    • pp.1053-1061
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    • 2004
  • Of the 20 arbitrarily chosen primers, six oligonucleotides decamer primers were used on the basis of the number of the polymorphisms generated in catfish (Silurus asotus) from Yesan and bullhead (Pseudobagrus fulvidraco) from Dangjin in Korea. Six primers were used generating a total of 602 scorable bands in catfish and 195 in bullhead population, respectively, ranging in size of DNA fragments from less than approximately 100 to larger than 2,000 base pairs (bp). Six primers yielded 199 polymorphic fragments (33.1%) in catfish and 47 (24%) in bullhead, respectively. In the present study, a total of 328 common fragments (an average of 54.7 per primer) were observed in catfish population, whereas 84 (an average of 14.0 per primer) in bullhead. The total number of specific fragments in catfish and bullhead population were 76 and 64, respectively. In catfish population, random decamer, OPA-17 (GACCGCTTGT) generated the highest number of fragments (a total of 141) in comparison with other primers used, with an average of 11.8. The common bands in the molecular weight of 300 bp generated by random primer OPA-06 (GGTCCCTGAC) were present in every individuals in bullhead population. The major polymorphic bands in the molecular weight of 100 bp generated by OPA-17 were identified in lane 14, 15, 17, 18, 19 20 and 21, which were identifying species in bullhead population. The average bandsharing values (BS values) of all of the samples within catfish population ranged from 0.575 to 0.945, whereas 0.063-1.000 within bullhead population. The bandsharing value (index of similarity between individuals) between individual No. 5 and No. 9 showed the highest level within catfish population, whereas the bandsharing value between individual No. 1 and No. 2 showed the lowest level. The single linkage cluster analysis resulted from four primers, indicating four genetic groupings composed of group 1 (C1-C10, all of the catfish samples), group 2 (B11, B12, B13, B14, B16, B17, B18, B19), group 3 (B15) and group 4 (B20 and B21). The dendrogram reveals close relationships between individual identities within two species populations and individuals derived from the same ancestor, respectively. However, genetic distances between two species populations ranged from 0.124 to 0.333. The shortest genetic distance (0.042) displaying significant molecular differences was between individual No. 6 and No. 9 catfish population. The shortest genetic distance (0.033) displaying significant molecular differences also was between individual No. 18 and No. 19 in bullhead population. Reversely, the genetic distance of individual No. 20/21 among individuals in bullhead population was highest (0.333). This result showed that bullhead No. 20 and 21 were distinct from other individuals within bullhead population.

No Association of Hypoxia Inducible Factor-1α Gene Polymorphisms with Breast Cancer in North-West Indians

  • Sharma, Sarika;Kapahi, Ruhi;Sambyal, Vasudha;Guleria, Kamlesh;Manjari, Mridu;Sudan, Meena;Uppal, Manjit Singh;Singh, Neeti Rajan
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.22
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    • pp.9973-9978
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    • 2014
  • Background: Hypoxia inducible factor-1 alpha (HIF-$1{\alpha}$) is the key regulator of cellular responses to hypoxia and plays a central role in tumour growth. Presence of Single nucleotide polymorphisms (SNPs) in the critical regulatory domains of HIF-$1{\alpha}$ may result in the overexpression of the protein and subsequent changes in the expression of the downstream target genes. The aim of study was to investigate the association of three SNPs (g.C111A, g.C1772T and g.G1790A) of HIF-$1{\alpha}$ with the risk of breast cancer in North Indian sporadic breast cancer patients. Materials and Methods: A total of 400 subjects, including 200 healthy controls and 200 patients with breast cancer were recruited in this study. Genotypes were determined using polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) method. Results: The CC and CA genotype frequency of HIF-$1{\alpha}$ g.C111A polymorphism was 100 vs 99% and 0 vs 1% in breast cancer patients and healthy controls respectively. The frequencies of CC, CT and TT genotype of g.C1772T polymorphism were 76 vs 74.5%, 19 vs 21% and 5 vs 4.5% in breast cancer patients and control individuals respectively. There was no significant difference in genotype and allele frequencies of HIF-$1{\alpha}$ g.C1772T polymorphism between cases and control individuals (p>0.05). For g.G1790A genotypes, all patients and controls had only GG genotype. Conclusions: The three HIF-$1{\alpha}$ polymorphisms (g.C111A, g.C1772T and g.G1790A) are not associated with breast cancer risk in North-West Indian patients.