RESUMEN
A large number of plant RNA viruses circulate between plants and insects. For RNA viruses, host alternations may impose a differential selective pressure on viral populations and induce variations in viral genomes. Here, we report the variations in the 3'-terminal regions of the multiple-segment RNA virus Rice stripe virus (RSV) that were discovered through de novo assembly of the genome using RNA sequencing data from infected host plants and vector insects. The newly assembled RSV genome contained 16- and 15-nt extensions at the 3'-termini of two genome segments compared with the published reference RSV genome. Our study demonstrated that these extensional sequences were consistently observed in two RSV isolates belonging to distinct genetic subtypes in RSV-infected rice, wheat and tobacco. Moreover, the de novo assembled genome of Southern rice black-streaked dwarf virus also contained 3'-terminal extensions in five RNA segments compared with the reference genome. Time course experiments confirmed that the 3'-terminal extensions of RSV were enriched in the vector insects, were gradually eliminated in the host plant and potentially affected viral replication. These findings indicate that variations in the 3'-termini of viral genomes may be different adaptive strategies for plant RNA viruses in insects and plants.
Asunto(s)
Variación Genética , Genoma Viral , Interacciones Huésped-Patógeno/genética , Insectos Vectores/virología , Oryza/virología , Tenuivirus/genética , Animales , Secuencia de Bases , Nucleótidos/genética , Enfermedades de las Plantas/virología , Reoviridae/genética , Ribonucleoproteínas/metabolismo , Ribonucleoproteínas/ultraestructura , Tenuivirus/aislamiento & purificación , Tenuivirus/ultraestructura , Replicación Viral/genéticaRESUMEN
Uno de los elementos imprescindibles en la ingeniería genética de plantas es un sistema de selección eficiente. El propósito de este trabajo fue evaluar la sensibilidad al marcador de selección higromicina B, de callos embriogénicos obtenidos a partir del escutelo de semilla de tres variedades colombianas de arroz (FEDEARROZ 2000, FEDEARROZ 50 y FEDEARROZ 369). Además, se validó la respuesta de estas variedades al protocolo de regeneración empleado. Se probaron cuatro concentraciones del antibiótico (25 mg/L, 50 mg/L, 75 mg/L y 100 mg/L) más un control sin higromicina B. Los resultados obtenidos mostraron que una concentración de 50 mg/L de antibiótico en el medio de regeneración es adecuada para la selección. Con esta concentración se impide la formación de brotes, aunque los callos no mueren completamente. Por otra parte, se estableció que el protocolo de regeneración utilizado es de baja eficiencia y, por consiguiente, es necesario optimizarlo para poder usarlo en procesos de ingeniería genética de cultivares colombianos de arroz.
An efficient selection system is one of the most important elements of plant genetic engineering. The purpose of this study was to evaluate the sensitivity of scutellum-derived embriogenic calli obtained from three colombian rice varieties (FEDEARROZ 2000, FEDEARROZ 50 and FEDEARROZ 369), to the selection marker hygromycin B. Aditionally, the response of these varieties to the regeneration protocol was measured. Four antibiotic concentrations were tested (25 mg/L, 50 mg/L, 75 mg/L and 100 mg/L) plus one control without hygromycin B. The results show that 50 mg/L of antibiotic in the regeneration medium is adequate for selection. This concentration prevents the formation of shoots, though the calli do not die. It was also established that the regeneration protocol is a low-efficiency system and it needs to be improved, in order to use it for colombian rice genetic engineering.
