RESUMEN
We report the successfull delay of leaf senescence in Medicago sativa. A highly regenerable clone of alfalfa was transformed with the construct SAG12-IPT, an approach that has already proved efficient in other crops. Several independent transformants were obtained as determined by Southern analysis and all the transformants expressed the transgene as measured by RT-PCR. In vitro and in vivo analyses showed that SAG12-IPT plants exhibited a stay-green phenotype that has the potential to greatly improve the quantity and quality of alfalfa forage.
Asunto(s)
Transferasas Alquil y Aril/genética , Senescencia Celular , Medicago sativa/fisiología , Hojas de la Planta/fisiología , Regiones Promotoras Genéticas/genética , Transformación Genética , Transferasas Alquil y Aril/metabolismo , Southern Blotting , Clorofila/metabolismo , Regulación Enzimológica de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Medicago sativa/genética , Fenotipo , Hojas de la Planta/enzimología , Plantas Modificadas Genéticamente , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The effects of increasing light and of a heterologous bHLH transcription factor on the accumulation of condensed tannins (CT) were investigated in leaves of Lotus corniculatus, a model legume species which accumulates these secondary metabolites in leaves as well as reproductive tissues. Light and expression of the transgene increased the level of CT in a synergistic way. To monitor how the changes in accumulation of condensed tannins were achieved, the level of expression of four key genes in the flavonoid pathway was estimated by real-time RT-PCR analysis. Early genes of the pathway (PAL and CHS) were affected less in their expression and so appeared to be less involved in influencing the final level of CT than later genes in the pathway (DFR and ANS). Steady-state levels of DFR and ANS transcripts showed a strong positive correlation with CT and these genes might be considered the first rate-limiting steps in CT biosynthesis in Lotus leaves. However, additional factors mediated by light are limiting CT accumulation once these genes are up-regulated by the transgene. Therefore, the increment of the steady-state mRNA level for DFR and ANS might not be sufficient to up-regulate condensed tannins in leaves. The real-time RT-PCR approach adopted showed that members within the CHS and DFR gene families are differentially regulated by the exogenous bHLH gene and light. This finding is discussed in relation to the approaches for controlling CT biosynthesis and for studying the expression profile of multi-gene families.