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Open access

Tomasz Oszako, Katarzyna Sikora, Lassaâd Belbahri and Justyna A. Nowakowska

Abstract

In Poland, about 20% of forest nurseries use irrigation water coming from natural superficial reservoirs, presumed to be the first source of infection caused by harmful pathogens belonging to the Oomycota class, especially Phytophthora genus and Pythium genus. The forest nursery is the only place where forest managers can react before pathogens leave it with asymptomatic plants or soil attached to their roots. The aim of this research was detection and identification phytopathogens in water samples. In order to recognise genus Phytophthora or Pythium in water collected from 33 places in five different forest districts in Poland, two DNA-based approaches of identification were applied: (i) the TaqMan probes, and (ii) sequencing of the ITS6/4 region.

The genomic DNA was obtained from 17 of 33 investigated water samples. TaqMan probes helped to identify 8 oomycetes present in 17 water samples. Based on ITS rDNA sequencing data, pathogens were identified in 17 cases, and this to the genus level (6 cases) and to the species level (11 cases). In total five Oomycetes species were identified, i.e. 3 Pythium species (Py. citrinum, Py. angustatum, Py. helicoides) and two Phytophthora species (P. lacustris sp. nov. - former taxon Salixsoil, P. gallica sp. nov.).

Open access

Agnieszka Kędrak-Jabłońska, Sylwia Budniak, Marek Krupa, Anna Szczawińska, Monika Reksa, Krzysztof Szulowski and Wojciech Iwaniak

References 1. Amagliani G., Omiccioli E., Brandi G., Bruce I.J., Magnani M.: A multiplex magnetic capture hybridisation and multiplex real-time PCR protocol for pathogen detection in sea food. Food Microbiol 2010, 27, 580–585. 2. Barkallah M., Gharbi Y., Hmani M., Mallek Z., Gautier M., Gdoura R., Fendri I.: Locked nucleic acid probe-based real-time PCR for the diagnosis of Listeria monocytogenes in ruminants. Mol Cell Probe 2016, 30, 138–145. 3. Budniak S., Kędrak-Jabłońska A., Szczawińska A., Reksa M., Krupa M., Szulowski K.: Comparison of

Open access

Agnieszka Kędrak-Jabłońska, Sylwia Budniak, Anna Szczawińska, Monika Reksa, Marek Krupa and Krzysztof Szulowski

References 1. Barbau-Piednoir E., Botteldoorn N., Yde M., Mahillon J., Roosens N.H.: Development and validation of qualitative SYBR®Green Real-Time PCR for detection and discrimination of Listeria spp. and Listeria monocytogenes. Appl Microbiol Biotechnol 2013, 97, 4021-4037. 2. Bassler H.A., Flood S.J.A., Livak K.J., Marmaro J., Knorr R., Batt C.A.: Use of a fluorogenic probe in a PCR-based assay for the detection of Listeria monocytogenes. Appl Environ Microbiol 1995, 61, 3724-3728. 3. Dmowska K., Wieczorek K

Open access

Małgorzata Natonek-Wiśniewska and Piotr Krzyścin

. (2004). A multiplex PCR assay for the identification of animal species in feedstuffs. Mol. Cell. Probe., 18: 81–87. Dooley J., Paine K., Garrett S., Brown H. (2004). Detection of meat species using TaqMan real-time PCR assays. Meat Sci., 68: 431–438. Farag M., Alagawany M., Abd El-Hack M., Tiwari R., Dhama K. (2015). Identification of different animal species in meat and meat products: trends and advances. Adv. Anim. Vet. Sci., 3: 334–346. Frezza D., Favaro M., Vaccari G., Holst C., Giambra V., Anklam E. (2003). A competitive polymerase chain

Open access

Yi-Xuan Hou, Chun Xie, Kang Wang, Yu-Ting Zhao, Yang-Yang Xie, Hong-Yan Shi, Jian-Fei Chen, Li Feng, Guang-Zhi Tong, Xiu-Guo Hua, Cong-Li Yuan, Yan-Jun Zhou and Zhi-Biao Yang

amplification for rapid detection of porcine epidemic diarrhea virus. Virus Genes 2011, 42, 229–235. 21. Salin C., Thaweesak C., Mai L Q., Sudarat D., Arunee C., Alongkorn A., Olfert L., Thaweesak S., Apiradee T.B., Yong P.R. H5N1 Oseltamivir-resistance detection by real-time PCR using two high sensitivity labeled TaqMan probes. J Virol Methods 2007, 139, 44–49. 22. Song D., Park B.: Porcine epidemic diarrhoea virus: a comprehensive review of molecular epidemiology, diagnosis, and vaccines. Virus genes 2012, 44, 167–175. 23. Sun R.Q, Cai R.J., Chen Y

Open access

Artur Jabłoński, Dominika Borowska, Sylwia Zębek, Andrzej Kowalczyk, Arkadiusz Dors, Jacek Żmudzki, Agnieszka Nowak and Zygmunt Pejsak

erythrocyte niche. PLoS One 6, 19574. 4. Guimaraes A.M., Vieira R.F., Poletto R., Vemulapalli R., Santos A.P., de Moraes W., Cubas Z.S., Santos L.C., Marchant- Forde J.N., Timenetsky J., Biondo A.W., Messick J.B.: A quantitative TaqMan PCR assay for the detection of Mycoplasma suis. J Appl Microbiol 2011, 111, 417-425. 5. Gwaltney S.M., Hays M.P., Oberst R.D.: Detection of Eperythrozoon suis using the polymerase chain reaction. J Vet Diagn Invest 1993, 5, 40-46. 6. Henderson J.P., O'Hagan J., Hawe S.M., Pratt M.C.: Anaemia and

Open access

Piotr Gronek, Joanna Gronek, Ewelina Lulińska-Kuklik, Michał Spieszny, Marta Niewczas, Mariusz Kaczmarczyk, Miroslav Petr, Patricia Fischerova, Ildus I. Ahmetov and Piotr Żmijewski

genotyped in duplicate. NOS3 , UCP2 , and AMPD1 genotyping The samples were genotyped using an allelic discrimination assay with a C1000 Touch Thermal Cycler (Bio-Rad, Germany) instrument with TaqMan® probes. To discriminate NOS3 Glu298Asp (NP_000594.2:p.Asp298Glu), UCP2 Ala55Val (NP_003346.2:p.Ala55Val), and AMPD1 Gln45Ter (NP_000027.2:p.Gln45Ter) alleles, TaqMan® Pre-Designed SNP Genotyping Assays were used (Applied Biosystems, USA) (assay ID: C_590093_1, C_3219460_20, C_903746_1, and C_33603912_10 respectively), including primers and fluorescently labelled

Open access

Monika Kałużna, Joanna Puławska and Artur Mikiciński

ABSTRACT

Phytosanitary measures to control diseases require sensitive and reliable detection methods. In case of Erwinia amylovora, the causal agent of fire blight, there are several methods developed. We tested six of them: (1) a conventional method; (2) standard PCR; (3) BIO-PCR; (4) Real-time PCR with SYBR Green; (5) Real-time PCR with TaqMan probe; and (6) Loop Mediated Isothermal Amplification (LAMP) in terms of their specificity, sensitivity and repeatability. Of the methods tested, only Real-time PCR with SYBR Green amplified non-specific fragments, which could be however identified by melting curve analysis. Real-time PCRs (both with SYBR Green and TaqMan probe) and LAMP were most sensitive and allowed to detect E. amylovora even in the samples where the concentration of cells was lower than 2 x 101 cfu/reaction. The highest repeatability was obtained using conventional method and Real-time PCRs. Of the methods used, only the LAMP technique was very insensitive to plant impurities.

Open access

Katarzyna Łosiewicz, Małgorzata Chmielewska-Krzesińska, Piotr Socha, Anna Jakimiuk and Krzysztof Wąsowicz

Abstract

The expression of miRNA-21, miRNA-10b, and miRNA-34a in malignant and benign tumours and non-neoplastic lesions in canine mammary gland, using real-time PCR with TaqMan probes was determined. The expression in normal tissues was compared to neoplastic and non-neoplastic lesions using one-way ANOVA test. Significant changes in miRNA expression in neoplastic tissues, as compared to normal ones, were demonstrated. In all neoplastic tissues, the miRNA-21 expression increased while in non-neoplastic lesions slightly decreased in comparison to normal ones. MiRNA-10b expression in malignant and benign tumours increased in comparison to normal tissues and non-neoplastic lesions. MiRNA-34a expression profile in neoplastic and non-neoplastic tissues differed from other examined miRNAs (miRNA-21 and miRNA-10b). In all samples miRNA-34a expression level decreased in comparison to normal tissues.

Open access

Georgijs Moisejevs, Linda Gailīte, Sergejs Isajevs, Liene Ņikitina-Zaķe, Inga Kempa, Dainius Jančiauskas, Ilze Ķikuste, Armands Sīviņš, Guntis Ancāns and Mārcis Leja

Abstract

Histamine has an important role in the process of the gastric mucosa inflammation acting via histamine receptor H2 (encoded by the gene HRH2). Single nucleotide polymorphism of the enhancer element of HRH2 gene promoter rs2067474 (1018G>A)may be associated with changes of expression of the receptor. We attempted to clarify the association of this polymorphism with gastric cancer and/or atrophic gastritis in the Latvian (Caucasian) population. The study group consisted of 121 gastric cancer patients and 650 patients with no evidence of gastric neoplasia on upper gastrointestinal endoscopy. Genotyping for rs2067474 was performed with the TaqMan probe-based system using a commercially available probe for RT-PCR. The frequency of the A allele in the gastric cancer group was 0.41% and in the control group — 1.54% (p = 0.231). No significant differences were found comparing genotypes between gastric cancer versus control patients (OR = 0.236, CI95% = 0.030–1.896), patients with (n = 165) versus without (n = 485) gastric metaplastic lesions (OR = 0.854, CI95% = 0.288–2.540) and patients with (n = 297) and without (n = 353) gastric atrophic lesions (OR = 1.145, CI95% = 0.451–2.906). Our findings suggest that the HRH2 -1018G>A polymorphism (rs2067474) is neither associated with gastric cancer nor the grade of atrophic gastritis in the Latvian (Caucasian) population.