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Food Control ◽  
2022 ◽  
Vol 132 ◽  
pp. 108494
Author(s):  
Sarah Azinheiro ◽  
Foteini Roumani ◽  
Laura Rodríguez-Lorenzo ◽  
Joana Carvalho ◽  
Marta Prado ◽  
...  

2022 ◽  
Vol 8 (1) ◽  
Author(s):  
Efthymia Stamelou ◽  
Ioannis A. Giantsis ◽  
Konstantinos V. Papageorgiou ◽  
Evanthia Petridou ◽  
Irit Davidson ◽  
...  

Abstract Backround Astrovirus, Norovirus and Sapovirus exhibit a wide distribution in swine pig herds worldwide. However, the association of porcine Astrovirus (PAstV), porcine Norovirus (PoNoV) and porcine Sapovirus (PoSaV) with disease in pigs remains uncertain. In this study, we investigated the prevalence of PAstV, PoNoV and PoSaV in Greek pig farms using both conventional RT-PCR and SYBR-Green Real-time RT-PCR in an effort to compare the sensitivity of the two methods. We examined 1400 stool samples of asymptomatic pigs originating from 28 swine farms throughout Greece in pools of five. Results PAstV was detected in all 28 swine farms examined, with an overall prevalence of 267/280 positive pools (95.4%). Porcine Caliciviruses prevalence was found at 36 and 57 out of the 280 examined samples, by the conventional and SYBR-Green Real time RT-PCR, respectively. Sequencing and phylogenetic analysis of the positive samples revealed that the detected PAstV sequences are clustered within PAstV1, 3 and 4 lineages, with PAstV3 being the predominant haplotype (91.2%). Interestingly, sequencing of the Calicivirus positive samples demonstrated the presence of non-target viruses, i.e. Sapovirus, Kobuvirus and Sapelovirus sequences and one sequence highly similar to bat Astrovirus, while no Norovirus sequence was detected. Conclusions The high prevalence of PAstV in Greek pig farms poses a necessity for further investigation of the pathogenicity of this virus and its inclusion in surveillance programs in case that it proves to be important. To our knowledge, this is the first epidemiological study of these viruses in pig farms in Greece.


2022 ◽  
Author(s):  
Vemula Harshini ◽  
S.M.K. Karthickeyan ◽  
K.G P. Kumarasamy ◽  
Tirumurugaan ◽  
C. Jeevan

Abstract A SYBR green real-time PCR assay was developed to find out the sex skewness in bovine sex-sorted semen samples. The qPCR assay of PLP and SRY genes revealed the mean values of X- and Y-bearing spermatozoa as 50.24 ± 0.65 and 49.75 ± 0.62 per cent in unsorted, and 91.80 ± 0.79 and 8.20 ± 0.73 per cent in X-enriched semen samples respectively.. The amplification efficiencies of the PLP and SRY primers were 99.25 and 98.03 per cent respectively. The method was validated by a series of repeatability and reproducibility assays which revealed low co-efficients of variations as 2.19 and 3.12 per cent respectively Thus becoming a reliable and inexpensive tool to evaluate the sorted semen on routine basis and validation of other sperm sexing technologies.


Virus Genes ◽  
2022 ◽  
Author(s):  
Jitendra K. Biswal ◽  
Biswa Ranjan Jena ◽  
Syed Zeeshan Ali ◽  
Rajeev Ranjan ◽  
Jajati K. Mohapatra ◽  
...  

Author(s):  
Fadi Abdel Sater ◽  
Mahmoud Younes ◽  
Hassan Nassar ◽  
Paul Nguewa ◽  
Kassem Hamze
Keyword(s):  
Low Cost ◽  

2021 ◽  
Vol 32 (6) ◽  
pp. e21682
Author(s):  
Hugo Gonzales Figueroa ◽  
Yat Sen Wong ◽  
Hugo Mauricio Gonzales Molfino ◽  
José Luis Llanos Carrillo

Se analizaron las propiedades biológicas de las células madre mesenquimales (CMM) provenientes del líquido sinovial de las articulaciones de las extremidades de caballo. Las muestras fueron caracterizadas por su peculiaridad de adherirse al plástico de los frascos de cultivos. Se utilizó la técnica de campos electromagnéticos (MACS) y el biomarcador de superficie STRO-1 con la finalidad de optimizar su aislamiento mediante el enriquecimiento de células precursoras mesenquimales (CMM-LS). Para determinar la expresión de los genes multipotencia, se extrajo el ARN total y se usó la transcriptasa reversa M-MLV para el ADNc, el que sirvió como molde para la qPCR (PCR cuantitativa). El qPCR se realizó utilizando el sistema BioRad con cuantificación mediante Sybr Green. Los genes de multipotencia cuantificados fueron OCT4 y NANOG. Las células seleccionadas utilizando MACS con el biomarcador de superficie STRO-1 (CMM-LS STRO-1), proliferan más rápidamente que las CMM-LS obtenidas sin purificar. Además, manifestaron una mayor predisposición a diferenciarse en condrocitos y osteocitos en comparación a las CMM-LS sin purificar, y una menor predisposición a diferenciarse en adipocitos. Así mismo, la expresión de OCT-4 fue significativamente superior en la CMM-LS STRO-1 en comparación con las CMM-LS sin purificar. La expresión de NANOG fue ligeramente superior en las CMM-LS STRO-1, pero sin diferencia significativa. Los resultados hacen suponer que la coexpresión de OCT-4 y Nanog podrían incrementar las funciones fisiológicas de las CMM-LS STRO-1 relacionadas con una mejor proliferación, autorrenovación y la predisposición a diferenciarse en los linajes osteocondrales; sin embargo, se necesita aclarar el papel funcional de estos marcadores de pluripotencia en células madre adultas.


2021 ◽  
Vol 9 (12) ◽  
pp. 2627
Author(s):  
María Paz Peris ◽  
Adriana Esteban-Gil ◽  
Paula Ortega-Hernández ◽  
Mariano Morales ◽  
Nabil Halaihel ◽  
...  

Canine leishmaniasis (CanL) diagnosis is not fully resolved. Currently, two specific methodologies are in continuous development, the detection of the parasite DNA or RNA in target organs and the detection of specific antibodies against Leishmania sp. For a correct diagnosis, it has been shown that the joint use of this type of test is necessary. In this work, a Sybr Green and a TaqMan Probe based on real time PCRs (qPCR) was performed for the detection of Leishmania sp. in order to correlate the results with clinicopathological and serological evaluations (IFA, ELISA and DAT) to propose an optimal biological sample to be used to detect the parasite in both early and late stages of the infection. A total of four samples were processed: conjunctival swabs, popliteal lymph node aspirates, bone marrow aspirates, and peripheral blood from experimentally infected dogs belonging to a larger study. Our results indicated that a single non-invasive sample (conjunctival swab) and the application of both types of qPCR would be reliable for determining Leishmania infection as well as the disease stage in dogs, thus avoiding bone marrow, lymph node aspirate or blood samples collection.


2021 ◽  
Vol 12 ◽  
Author(s):  
Lais Sevilha-Santos ◽  
Selma Regina Penha Silva Cerqueira ◽  
Ciro Martins Gomes

The development of new molecular techniques is essential for the early diagnosis of leprosy. Studies in the field have failed to elucidate the performance of these tests in clinical practice. We aimed to design a new primer pair for the repetitive element (RLEP) target of Mycobacterium leprae and to test the accuracy of SYBR green-based real-time PCR through the evaluation of different thresholds for different skin layers. We also aimed to track the transmission potential of multibacillary and paucibacillary leprosy patients. The in vitro validation of our reaction resulted in a quantification limit of 0.03 bacilli. We then conducted a cross-sectional/cohort-based study of diagnostic accuracy. Patients were included, and skin samples were divided into four layers: epidermis, superior dermis, inferior dermis, and hypodermis. We also quantified M. leprae in nasal swabs of the included patients and compared the results to the number of household contacts also diagnosed with leprosy. One hundred patients with a clinical presentation compatible with leprosy were allocated to the leprosy or control group. Although the parasite load was greater in the superior and inferior dermis, M. leprae DNA was found in all skin layers. The best sensitivity was observed for the superior dermis using the presence of any quantifiable bacillus DNA as the threshold [sensitivity=59.26% (95% CI=45.97–71.32)]. In the epidermis, setting 1 quantifiable bacillus as the threshold resulted in 100% specificity (95% CI=92.29–100). The number of bacilli found in nasal swabs was not significantly related to the number of household contacts also diagnosed with leprosy. Paucibacillary patients tested positive only for bacillus fragments in nasal swabs but not for the entire bacilli. We can conclude that superficial biopsies might result in sensitivity loss, although different skin sample types will have little influence on the final accuracy. In contrast, threshold changes greatly influence these properties. Paucibacillary patients may not be a relevant source of disease transmission.


2021 ◽  
Vol 1 ◽  
Author(s):  
Shafariatul Akmar-Ishak ◽  
Fifi Fariza-Azmi ◽  
Adibah Syahnaz-Zahari ◽  
Dayang Fredalina-Basri

Background: Malaria is a parasite that is transmitted to human through the bite of a female Anopheles mosquito. Every year human was exposed to the threat of malaria infection. This disease becomes more fatal as these parasites show resistance towards the drug available. Thus, searches for new antimalarial drug are crucial. This study was carried out to evaluate the antimalarial activity in Canarium odontophyllum leaf extracts (methanol, acetone and aqueous) against erythrocytes infected with Plasmodium berghei NK65 using Plasmodium Lactate Dehydrogenase (pLDH) Assay and SYBR green I fluorescence Assay. Method: Three types of solvents were used to extracts Canarium odontophyllum leaf according to increasing polarity index; acetone, methanol and aqueous. These extracts were made into eight-fold serial dilution; concentrations ranging from 0.00001μg / ml as the lowest concentration until 100 μg / ml as the highest concentration and further tested on Plasmodium berghei NK65 infected erythrocytes via ex-vivo. The IC50 (inhibition concentration) 50 readings were taken at the point of 5% parasitemia level and in the synchronization process. Both PLDH assay and SYBR green I fluorescence assay were being carried out simultaneously. Result: The One-way ANOVA showed that there is no significant difference between extracts at 5% parasitemia level, even so methanol was further tested on synchronization process as it showed the lowest reading of IC50 among the three extracts for PLDH assay and SYBR green I fluorescence assay respectively, (IC50 0.00045μg / ml, 0.002 μg / ml). For synchronization stages, the One-way ANOVA result showed there is no significant difference between stages of morphology. However, methanol extracts showed the most potent on schizont, (1.16x10-5 μg / ml ) and young trophozoite, (0.00195 μg / ml ) stages for each method respectively. Conclusions: All three extracts of Canarium odontophyllum leaf were effective on Plasmodium berghei NK65, however methanol showed most promising results and further research on the fractions were required for proper drug development. The Ministry of Higher Learning funded this project, Government of Malaysia, under the Fundamental Research Grant Scheme Code No. FRGS/2/2014/SG05/UKM/02/3.


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