Quantitative PCR for detection of Citrus tristeza virus in Colombia
Real-time quantitative PCR (qRT-PCR) was applied using SYBR Green for the specific detection of Citrus tristeza virus (CTV) in Colombian. Genomic RNA (gRNA) was amplified using primers designed from conserved sequences in the open reading frames (ORF’s) 1b and 2. We obtained a 186 bp product with ne...
| Main Authors: | , , |
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| Format: | Online |
| Language: | spa |
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Universidad Pedagógica y Tecnológica de Colombia
2018
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| Subjects: | |
| Online Access: | https://revistas.uptc.edu.co/index.php/ciencia_agricultura/article/view/7789 |
| _version_ | 1801706474108354560 |
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| author | Solano-Luna, Luis Miguel Chavarro-Mesa, Edisson Ángel-Diaz, Jorge Evelio |
| author_facet | Solano-Luna, Luis Miguel Chavarro-Mesa, Edisson Ángel-Diaz, Jorge Evelio |
| author_sort | Solano-Luna, Luis Miguel |
| collection | OJS |
| description | Real-time quantitative PCR (qRT-PCR) was applied using SYBR Green for the specific detection of Citrus tristeza virus (CTV) in Colombian. Genomic RNA (gRNA) was amplified using primers designed from conserved sequences in the open reading frames (ORF’s) 1b and 2. We obtained a 186 bp product with neither dimers nor non-specificity. The analysis of the melting curve showed a peak between 81 °C and 83 °C, with a correlation coefficient of 0.998 and an efficiency of 99.1 %. The amplification of the 186 bp fragment resulted in the standard curve, which allowed the quantitative analysis of the samples with a detection range between 1x108 and 1x103 genomic RNA copies, with low variation coefficients. CTV accumulation was higher in foliar and fruit tissue than in the bark, and the differences observed among several citrus species susceptible to infection were minimal. The highest concentration of virus was found in the upper third of the analyzed plants, followed by the lower third and finally the middle third. The qRT-PCR is a specific and sensitive method, with a practical interest for the detection of viral diseases in citrus plants and other crops of commercial interest. |
| format | Online |
| id | oai:oai.revistas.uptc.edu.co:article-7789 |
| institution | Revista Ciencia y Agricultura |
| language | spa |
| publishDate | 2018 |
| publisher | Universidad Pedagógica y Tecnológica de Colombia |
| record_format | ojs |
| spelling | oai:oai.revistas.uptc.edu.co:article-77892022-06-16T15:19:36Z Quantitative PCR for detection of Citrus tristeza virus in Colombia PCR cuantitativa para la detección del virus de la tristeza de los cítricos en Colombia Solano-Luna, Luis Miguel Chavarro-Mesa, Edisson Ángel-Diaz, Jorge Evelio absolute quantification citrus diseases Closterovirus detection of plant pathogens real-Time PCR SYBR Green Closterovirus cuantificación absoluta detección de patógenos vegetales enfermedades de los cítricos PCR en tiempo real SYBR Green Real-time quantitative PCR (qRT-PCR) was applied using SYBR Green for the specific detection of Citrus tristeza virus (CTV) in Colombian. Genomic RNA (gRNA) was amplified using primers designed from conserved sequences in the open reading frames (ORF’s) 1b and 2. We obtained a 186 bp product with neither dimers nor non-specificity. The analysis of the melting curve showed a peak between 81 °C and 83 °C, with a correlation coefficient of 0.998 and an efficiency of 99.1 %. The amplification of the 186 bp fragment resulted in the standard curve, which allowed the quantitative analysis of the samples with a detection range between 1x108 and 1x103 genomic RNA copies, with low variation coefficients. CTV accumulation was higher in foliar and fruit tissue than in the bark, and the differences observed among several citrus species susceptible to infection were minimal. The highest concentration of virus was found in the upper third of the analyzed plants, followed by the lower third and finally the middle third. The qRT-PCR is a specific and sensitive method, with a practical interest for the detection of viral diseases in citrus plants and other crops of commercial interest. Se aplicó la técnica de reacción en cadena de la polimerasa en tiempo real (qRT-PCR), usando SYBR Green para la detección específica del virus de la tristeza de los cítricos (CTV) en Colombia. Una pareja de iniciadores, diseñados a partir de secuencias conservadas en los marcos abiertos de lectura (ORF’s) 1b y 2, permitió amplificar el ARN genómico (ARNg), y un producto de 186 pb fue obtenido sin la presencia de dímeros o inespecificidad. El análisis de la curva de fusión mostró un pico entre 81 oC y 83 oC, con un coeficiente de correlación de 0,998 y una eficiencia de 99,1 %. La curva estándar se desarrolló a partir de una amplificación del producto de 186 pb y permitió realizar un análisis cuantitativo de las muestras, con un rango de detección de 1x108 hasta 1x103 copias de RNAg y con valores de coeficiente de variación bajos. La acumulación de CTV fue más alta en tejido foliar y en frutos que en corteza; entretanto, las diferencias demostradas entre varias especies de cítricos susceptibles a la infección fueron mínimas. Los resultados de este trabajo muestran que la mayor concentración de virus se encontró en el tercio superior de las plantas analizadas, seguida por el tercio bajo y, por último, el tercio medio. La qRT-PCR es un método específico y sensible de interés práctico en los procesos de detección de las enfermedades virales presentes en el cultivo de los cítricos y otros cultivos de interés comercial. Universidad Pedagógica y Tecnológica de Colombia 2018-02-01 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion research investigación application/pdf https://revistas.uptc.edu.co/index.php/ciencia_agricultura/article/view/7789 10.19053/01228420.v15.n1.2018.7789 Ciencia y Agricultura; Vol. 15 No. 1 (2018); 7-18 Ciencia y Agricultura; Vol. 15 Núm. 1 (2018); 7-18 2539-0899 spa https://revistas.uptc.edu.co/index.php/ciencia_agricultura/article/view/7789/6234 Copyright (c) 2018 Revista Ciencia y Agricultura |
| spellingShingle | absolute quantification citrus diseases Closterovirus detection of plant pathogens real-Time PCR SYBR Green Closterovirus cuantificación absoluta detección de patógenos vegetales enfermedades de los cítricos PCR en tiempo real SYBR Green Solano-Luna, Luis Miguel Chavarro-Mesa, Edisson Ángel-Diaz, Jorge Evelio Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title | Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title_alt | PCR cuantitativa para la detección del virus de la tristeza de los cítricos en Colombia |
| title_full | Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title_fullStr | Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title_full_unstemmed | Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title_short | Quantitative PCR for detection of Citrus tristeza virus in Colombia |
| title_sort | quantitative pcr for detection of citrus tristeza virus in colombia |
| topic | absolute quantification citrus diseases Closterovirus detection of plant pathogens real-Time PCR SYBR Green Closterovirus cuantificación absoluta detección de patógenos vegetales enfermedades de los cítricos PCR en tiempo real SYBR Green |
| topic_facet | absolute quantification citrus diseases Closterovirus detection of plant pathogens real-Time PCR SYBR Green Closterovirus cuantificación absoluta detección de patógenos vegetales enfermedades de los cítricos PCR en tiempo real SYBR Green |
| url | https://revistas.uptc.edu.co/index.php/ciencia_agricultura/article/view/7789 |
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