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Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control
Published on: March 30, 2015
El control positivo del empalme pre-ARNm en vitro es el control positivo del empalme pre-ARNm en vitro
1Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, MA 02138.
Science (New York, N.Y.)
|April 10, 1992
Resumen
Los genes transformador (tra) y transformador-2 (tra-2) controlan el empalme específico del sexo en Drosophila. Promueven un complejo proteico en un exón, facilitando el reconocimiento del sitio de empalme para el desarrollo femenino.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- Genética La genética.
- Biología del desarrollo Biología del desarrollo.
Sus antecedentes:
- El empalme alternativo regula la expresión génica, crucial para la diferenciación sexual.
- El empalme alternativo del gen de doble sexo (dsx) es clave para el desarrollo sexual en Drosophila melanogaster.
- Las proteínas del transformador (tra) y del transformador-2 (tra-2) son reguladores conocidos del empalme dsx.
Objetivo del estudio:
- Investigar el mecanismo in vitro mediante el cual el transformador (tra) y el transformador-2 (tra-2) activan el sitio de empalme 3' específico de la hembra del pre-ARNm.m de doble sexo (dsx).
- Identificar los elementos reguladores y las interacciones de proteínas involucradas en este evento de empalme alternativo específico del sexo.
Principales métodos:
- Utilizó un sistema de empalme in vitro para estudiar el procesamiento del pre-ARNm dsx.
- Las proteínas recombinantes del transformador (Tra) y del transformador-2 (Tra-2) se utilizaron para activar el sitio de empalme 3' específico de la hembra.
- Se realizaron ensayos de desplazamiento de movilidad electroforética (EMSA) para detectar la unión de proteínas a la secuencia de exones reguladoras.
Principales resultados:
- Se encontró que una secuencia de exones, esencial in vivo, era necesaria y suficiente para la activación in vitro del sitio de empalme 3' específico de la hembra.
- Las proteínas Tra y Tra-2 recombinantes podrían activar el sitio de empalme en el sistema in vitro.
- Se demostró la unión específica de proteínas nucleares, incluidas Tra y Tra-2, a la secuencia de exones reguladoras.
Conclusiones:
- Es probable que Tra y Tra-2 funcionen promoviendo el ensamblaje de un complejo multiproteico en una secuencia de exones específica del pre-ARNm dsx.
- Este complejo multiproteico puede mejorar el reconocimiento del sitio de empalme 3' adyacente específico de la hembra por la maquinaria de empalme.
- Los hallazgos proporcionan información mecanicista sobre la regulación del empalme alternativo específico por sexo en Drosophila.
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