Estandarización y optimización de técnicas moleculares para Aloe barbadensis Mill / Standardization and optimization of molecular techniques for Aloe barbadensis Mill.
Resumen
Resumen
La aceptación de protocolos de micropropagación de aloe (Aloe barbadensis Mill.) depende de la fidelidad genética de las plantas. Para estandarizar y optimizarlas condiciones de la PCR como técnica molecular, se extrajo ADN dela zona media-basal del tejido foliar de vitroplantas e hijuelos parentales utilizando el protocolo de 2% CTAB/1,4M NaCl y ARNasa. Las condiciones de la PCR evaluadas fueronla cantidad de ADN (1 a 100 ng) por reacción, la concentración del MgCl2 (1; 2; 3 y 4 mM) y la temperatura de hibridación (Ta) según la temperatura de melting (Tm) de 29 iniciadoresRAPD e ISSR. Se logró la estandarización y optimización para 18 iniciadores, destacando que los RAPD OPA 02, OPA 13, OPC 01, OPC 19 y OPC 20 son reportados por primera vez en aloe. Se evidenciaron18 iniciadoresRAPD e ISSRque podrán ser utilizados como marcadores molecularesen estudiosde fidelidad genéticaenplantas de aloe.
Abstract
The acceptance of micropropagation of aloe (Aloe barbadensis Mill.) depends on the genetic fidelity of the plants. In order to standardize and optimize the conditions of polymerize chain reaction (PCR)as molecular technique, DNA was extracted from intermediate-base zone of leaf tissue of parental offshoots and vitroplants, using 2% CTAB/1,4M NaCl and ARNasa protocol. PCR conditions evaluated were amount of DNA per reaction (1 to 100ng), concentration of MgCl2 (1; 2; 3 and 4 mM), and annealing temperature (Ta) according to the melting temperature (Tm) of 29 primersRAPD and ISSR. Standardization and optimization of PCR conditions was obtained for 18 primers, noting that the RAPD OPA 02, OPA 13, OPC 01, OPC 19andOPC 20 are the first report for aloe. There were 18 initiatorsRAPD e ISSR that can be used as molecular markers in genetic fidelity studies in aloe plants.
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