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Registro completo
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Biblioteca (s) : |
INIA Las Brujas. |
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Fecha : |
04/10/2014 |
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Actualizado : |
06/06/2023 |
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Tipo de producción científica : |
Artículos en Revistas Indexadas Internacionales |
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Autor : |
UMAÑA, R.; PRITSCH, C.; ARBIZA, J.R.; RIVAS, F.; PAGLIANO, G. |
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Afiliación : |
CARLOS FERNANDO RIVAS GRELA, INIA (Instituto Nacional de Investigación Agropecuaria), Uruguay. |
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Título : |
Evaluation of four viroid rna extraction methods for the molecular diagnosis of cevd in citrus lemon using rt-pcr, dot blot and northern blot. [Evaluación de cuatro métodos de extracción de ARN viroide para el diagnóstico molecular de CEVd en Citrus limon mediante RT-PCR, Dot blot y Northern blot.]. |
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Fecha de publicación : |
2013 |
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Fuente / Imprenta : |
Biotecnologia Aplicada, 2013, v.30, no.2, p.125-130. |
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Descripción física : |
2-s2.0-84884171392 |
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Serie : |
1027-2852 |
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ISSN : |
1027-2852 |
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Idioma : |
Inglés |
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Contenido : |
ABSTRACT.
An efficient method for RNA extraction that leads to RNA high yield and purity is a technical issue relevant for development and optimization of molecular diagnostic methods aimed to detect viroid infections in citrus varieties. Residual contaminants may affect RNA detection depending on the molecular diagnosis approaches. This condition can be evaluated through RNA absorption spectrum analysis. Functionally, it is assessed through observation of RTPCR amplification products and Northern blot and Dot-blot signal intensities, displaying levels of analytical response/ sensitivity. Four RNA extraction methods were evaluated to determine their effects on the capacity to detect viroid CEVd presence/absence in Citrus limon through four molecular diagnostic approaches: 1) conventional viroid extraction (CVE); 2) phenol/guanidine thiocyanate (PGT), 3) SDS/potassium acetate (SPA); and 4) formaldehyde/ SSC (FS). Phloem tissue quantifications showed values between 7500 ng/μL and 1200 ng/μL and ranged 1.3-2.0 OD260/280. Evaluations through RT-PCR showed the expected amplifications of the entire CEVd genome, but erratic scenarios still remained. Non-radioactive probe hybridization techniques revealed high intensity signals (132 RU) for infected tissue, by using the CVE method, and a positivity cut-off for the presence of infection was established (78 RU). Nevertheless, molecular hybridization tools can jeopardize the diagnosis due to the thoroughness of the protocol and the RNA template conditions. The diagnostic ability of the association of Northern blot with CVE viroid extraction
analyses as starting point was evidenced for successful detection, among the molecular methods tested.
RESUMEN.Evaluación de cuatro métodos de extracción de ARN viroide para el diagnóstico molecular de CEVd en Citrus limon mediante RT-PCR, Dot blot y Northern blot. En el desarrollo de metodologías moleculares diagnósticas en variedades cítricas propensas a infección viroide, se precisa la extracción efi ciente de ARN, siguiendo criterios de concentración y pureza. Los contaminantes remanentes pueden afectar la detección según la herramienta molecular escogida. Esta condición se analiza a partir del espectro de absorción del ARN; mediante la reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) y por los niveles de intensidad de las señales de Northern blot y Dot blot, en términos de respuesta analítica/sensibilidad. Se evaluaron cuatro métodos de extracción de ARN, por sus
efectos sobre la detección de presencia/ausencia del Citrus Exocortis Viroid (CEVd) en Citrus limon mediante análisis moleculares diagnósticos: 1) extracción viroide convencional (EVC); 2) fenol/tiocianato de guanidina (FTG); 3) SDS/ acetato de potasio (SAP); y 4) formaldehído/SSC (FS). Los valores del tejido de fl oema estuvieron entre 7500 ng/μL y 1200 ng/μL y los rangos entre 1.3 y 2.0 DO260/280. La evaluación por aproximaciones de la RT-PCR reflejó las amplifi caciones esperadas del genoma completo del CEVd; sin embargo, aún se discuten los escenarios erráticos.
Las hibridaciones no radiactivas revelaron señales de alta intensidad (132 UR) para el tejido infectado, según el método de EVC, y la defi nición del límite de positividad para la presencia de infección (78 UR). Las herramientas basadas en hibridaciones moleculares interfi eren en el diagnóstico, por la rigurosidad del protocolo y las condiciones del ARN molde. La extracción viroide como punto de partida de una detección exitosa y los métodos moleculares ensayados, mostraron las posibilidades diagnósticas de la asociación de Northern blot con la EVC. MenosABSTRACT.
An efficient method for RNA extraction that leads to RNA high yield and purity is a technical issue relevant for development and optimization of molecular diagnostic methods aimed to detect viroid infections in citrus varieties. Residual contaminants may affect RNA detection depending on the molecular diagnosis approaches. This condition can be evaluated through RNA absorption spectrum analysis. Functionally, it is assessed through observation of RTPCR amplification products and Northern blot and Dot-blot signal intensities, displaying levels of analytical response/ sensitivity. Four RNA extraction methods were evaluated to determine their effects on the capacity to detect viroid CEVd presence/absence in Citrus limon through four molecular diagnostic approaches: 1) conventional viroid extraction (CVE); 2) phenol/guanidine thiocyanate (PGT), 3) SDS/potassium acetate (SPA); and 4) formaldehyde/ SSC (FS). Phloem tissue quantifications showed values between 7500 ng/μL and 1200 ng/μL and ranged 1.3-2.0 OD260/280. Evaluations through RT-PCR showed the expected amplifications of the entire CEVd genome, but erratic scenarios still remained. Non-radioactive probe hybridization techniques revealed high intensity signals (132 RU) for infected tissue, by using the CVE method, and a positivity cut-off for the presence of infection was established (78 RU). Nevertheless, molecular hybridization tools can jeopardize the diagnosis due to the thoroughness of the protocol ... Presentar Todo |
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Palabras claves : |
CEVd. |
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Thesagro : |
CITRUS; DETECCIÓN DE VIROIDES DE CITRUS; DIAGNOSTICO; ENFERMEDADES DE LOS CITRUS; TECNICAS; VIROSIS. |
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Asunto categoría : |
--
H20 Enfermedades de las plantas |
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URL : |
http://www.ainfo.inia.uy/digital/bitstream/item/3438/1/Rivas-F.-2013.-Biotec.Aplicada-v.302-p.125-130.pdf
http://www.ainfo.inia.uy/digital/bitstream/item/3439/1/Rivas-F.-2013.-Biotec.Aplicada-v.302-p.131-136.pdf
http://www.medigraphic.com/pdfs/biotecapl/ba-2013/ba132g.pdf
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Marc : |
LEADER 04714naa a2200289 a 4500
001 1050903
005 2023-06-06
008 2013 bl uuuu u00u1 u #d
022 $a1027-2852
100 1 $aUMAÑA, R.
245 $aEvaluation of four viroid rna extraction methods for the molecular diagnosis of cevd in citrus lemon using rt-pcr, dot blot and northern blot. [Evaluación de cuatro métodos de extracción de ARN viroide para el diagnóstico molecular de CEVd en Citrus limon mediante RT-PCR, Dot blot y Northern blot.].$h[electronic resource]
260 $c2013
300 $c2-s2.0-84884171392
490 $a1027-2852
520 $aABSTRACT. An efficient method for RNA extraction that leads to RNA high yield and purity is a technical issue relevant for development and optimization of molecular diagnostic methods aimed to detect viroid infections in citrus varieties. Residual contaminants may affect RNA detection depending on the molecular diagnosis approaches. This condition can be evaluated through RNA absorption spectrum analysis. Functionally, it is assessed through observation of RTPCR amplification products and Northern blot and Dot-blot signal intensities, displaying levels of analytical response/ sensitivity. Four RNA extraction methods were evaluated to determine their effects on the capacity to detect viroid CEVd presence/absence in Citrus limon through four molecular diagnostic approaches: 1) conventional viroid extraction (CVE); 2) phenol/guanidine thiocyanate (PGT), 3) SDS/potassium acetate (SPA); and 4) formaldehyde/ SSC (FS). Phloem tissue quantifications showed values between 7500 ng/μL and 1200 ng/μL and ranged 1.3-2.0 OD260/280. Evaluations through RT-PCR showed the expected amplifications of the entire CEVd genome, but erratic scenarios still remained. Non-radioactive probe hybridization techniques revealed high intensity signals (132 RU) for infected tissue, by using the CVE method, and a positivity cut-off for the presence of infection was established (78 RU). Nevertheless, molecular hybridization tools can jeopardize the diagnosis due to the thoroughness of the protocol and the RNA template conditions. The diagnostic ability of the association of Northern blot with CVE viroid extraction analyses as starting point was evidenced for successful detection, among the molecular methods tested. RESUMEN.Evaluación de cuatro métodos de extracción de ARN viroide para el diagnóstico molecular de CEVd en Citrus limon mediante RT-PCR, Dot blot y Northern blot. En el desarrollo de metodologías moleculares diagnósticas en variedades cítricas propensas a infección viroide, se precisa la extracción efi ciente de ARN, siguiendo criterios de concentración y pureza. Los contaminantes remanentes pueden afectar la detección según la herramienta molecular escogida. Esta condición se analiza a partir del espectro de absorción del ARN; mediante la reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) y por los niveles de intensidad de las señales de Northern blot y Dot blot, en términos de respuesta analítica/sensibilidad. Se evaluaron cuatro métodos de extracción de ARN, por sus efectos sobre la detección de presencia/ausencia del Citrus Exocortis Viroid (CEVd) en Citrus limon mediante análisis moleculares diagnósticos: 1) extracción viroide convencional (EVC); 2) fenol/tiocianato de guanidina (FTG); 3) SDS/ acetato de potasio (SAP); y 4) formaldehído/SSC (FS). Los valores del tejido de fl oema estuvieron entre 7500 ng/μL y 1200 ng/μL y los rangos entre 1.3 y 2.0 DO260/280. La evaluación por aproximaciones de la RT-PCR reflejó las amplifi caciones esperadas del genoma completo del CEVd; sin embargo, aún se discuten los escenarios erráticos. Las hibridaciones no radiactivas revelaron señales de alta intensidad (132 UR) para el tejido infectado, según el método de EVC, y la defi nición del límite de positividad para la presencia de infección (78 UR). Las herramientas basadas en hibridaciones moleculares interfi eren en el diagnóstico, por la rigurosidad del protocolo y las condiciones del ARN molde. La extracción viroide como punto de partida de una detección exitosa y los métodos moleculares ensayados, mostraron las posibilidades diagnósticas de la asociación de Northern blot con la EVC.
650 $aCITRUS
650 $aDETECCIÓN DE VIROIDES DE CITRUS
650 $aDIAGNOSTICO
650 $aENFERMEDADES DE LOS CITRUS
650 $aTECNICAS
650 $aVIROSIS
653 $aCEVd
700 1 $aPRITSCH, C.
700 1 $aARBIZA, J.R.
700 1 $aRIVAS, F.
700 1 $aPAGLIANO, G.
773 $tBiotecnologia Aplicada, 2013$gv.30, no.2, p.125-130.
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Registro original : |
INIA Las Brujas (LB) |
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 | Acceso al texto completo restringido a Biblioteca INIA La Estanzuela. Por información adicional contacte bib_le@inia.org.uy. |
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Registro completo
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Biblioteca (s) : |
INIA La Estanzuela. |
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Fecha actual : |
08/09/2014 |
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Actualizado : |
07/11/2019 |
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Tipo de producción científica : |
Artículos en Revistas Indexadas Internacionales |
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Circulación / Nivel : |
A - 2 |
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Autor : |
CASTRO A.; GAMBA, F.; GERMÁN, S.; GONZÁLEZ, S.N.; HAYES, P.M.; PEREYRA, S.; PÉREZ, C. |
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Afiliación : |
SILVIA ELISA GERMAN FAEDO, Instituto Nacional de Investigación Agropecuaria (INIA), Uruguay; SILVANA NOEMÍ GONZÁLEZ PARODI, INIA La Estanzuela; SILVIA ANTONIA PEREYRA CORREA, Instituto Nacional de Investigación Agropecuaria (INIA), Uruguay. |
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Título : |
Quantitative trait locus analysis of spot blotch and leaf rust resistance in the BCD47 × Baronesse barley mapping population. |
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Fecha de publicación : |
2012 |
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Fuente / Imprenta : |
Plant Breeding, v. 131, n. 2, p. 258-266, 2012. |
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ISSN : |
0179-9541 |
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DOI : |
10.1111/j.1439-0523.2011.01930.x |
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Idioma : |
Inglés |
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Notas : |
Article histoty: Received December 22, 2010/Accepted October 26, 2011. |
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Contenido : |
Abstract:We studied the genetics of the resistance to leaf rust (LR) (caused by Puccinia hordei) and spot blotch (SB) (caused by Cochliobolus sativus)
in barley using a doubled-haploid population derived from the cross BCD47 · Baronesse. BCD47 has low SB severity and high susceptibility to LR, while Baronesse is susceptible to SB and has low LR severity. Resistance to both diseases is expressed at the adult plant stage. The population was phenotyped in eight field environments for SB and nine for LR. Ten quantitative trait loci (QTLs) were detected for SB. None were significant in more than three environments, and
both parents contributed resistance alleles. Five QTLs were detected for LR. The most consistent quantitative trait locus (QTL) (significant
in seven environments) was on chromosome 6H (located on the Bmag173-Bmag009 interval) with Baronesse contributing the resistance allele. Coincident QTL effects for SB were also detected in this region with resistance alleles to the two diseases in repulsion. These results illustrate the difficulties of resistance gene detection in the complex disease environments found under field conditions. |
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Palabras claves : |
COCHLIOBOLUS SATIVUS; DISEASE RESISTANCE; HORDEUM; LEAF RUST; PUCCINIA HORDEI; QUANTITATIVE TRAIT LOCI; SPOT BLOTCH. |
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Thesagro : |
FITOPATOLOGIA. |
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Asunto categoría : |
H20 Enfermedades de las plantas |
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Marc : |
LEADER 02163naa a2200325 a 4500
001 1050040
005 2019-11-07
008 2012 bl uuuu u00u1 u #d
022 $a0179-9541
024 7 $a10.1111/j.1439-0523.2011.01930.x$2DOI
100 1 $aCASTRO A.
245 $aQuantitative trait locus analysis of spot blotch and leaf rust resistance in the BCD47 × Baronesse barley mapping population.$h[electronic resource]
260 $c2012
500 $aArticle histoty: Received December 22, 2010/Accepted October 26, 2011.
520 $aAbstract:We studied the genetics of the resistance to leaf rust (LR) (caused by Puccinia hordei) and spot blotch (SB) (caused by Cochliobolus sativus) in barley using a doubled-haploid population derived from the cross BCD47 · Baronesse. BCD47 has low SB severity and high susceptibility to LR, while Baronesse is susceptible to SB and has low LR severity. Resistance to both diseases is expressed at the adult plant stage. The population was phenotyped in eight field environments for SB and nine for LR. Ten quantitative trait loci (QTLs) were detected for SB. None were significant in more than three environments, and both parents contributed resistance alleles. Five QTLs were detected for LR. The most consistent quantitative trait locus (QTL) (significant in seven environments) was on chromosome 6H (located on the Bmag173-Bmag009 interval) with Baronesse contributing the resistance allele. Coincident QTL effects for SB were also detected in this region with resistance alleles to the two diseases in repulsion. These results illustrate the difficulties of resistance gene detection in the complex disease environments found under field conditions.
650 $aFITOPATOLOGIA
653 $aCOCHLIOBOLUS SATIVUS
653 $aDISEASE RESISTANCE
653 $aHORDEUM
653 $aLEAF RUST
653 $aPUCCINIA HORDEI
653 $aQUANTITATIVE TRAIT LOCI
653 $aSPOT BLOTCH
700 1 $aGAMBA, F.
700 1 $aGERMÁN, S.
700 1 $aGONZÁLEZ, S.N.
700 1 $aHAYES, P.M.
700 1 $aPEREYRA, S.
700 1 $aPÉREZ, C.
773 $tPlant Breeding$gv. 131, n. 2, p. 258-266, 2012.
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