La Agarosa y sus usos en Biología Molecular
La Agarosa y sus usos en Biología Molecular
¿Qué es la Agarosa?
La agarosa es un polisacárido obtenido de algas rojas marinas, el cual está conformado por unidades de agarobiosa, un disacárido formado por las galactosas alfa y beta.
Es usada frecuentemente en biología molecular para la separación de moléculas por electroforesis, especialmente de ADN y ARN.
La agarosa se disuelve en caliente (50-60°C) y al enfriar solidifica formando un gel, de alta porosidad. Las cadenas de agarosa forman fibras helicoidales, que generan una malla tridimensional de canales con diámetros entre 50 y >200 nm, constituyendo una red fibrosa que puede ser utilizada como filtro para la separación de diferentes moléculas, siendo el tamaño del poro inversamente proporcional a la concentración de agarosa.
¿Quién descubrió el uso de la agarosa para la electroforesis de ADN?
La creación de la electroforesis como un método de separación de ADN bicatenario de cadenas simples se remonta al año 1962, elaborado por Matsubara y Takagi, cuando se utilizaba al almidón como gel de corrimiento.
La necesidad de analizar moléculas de ADN de longitudes variables propició el estudio de otros compuestos que pudieran utilizarse como gel de corrimiento en las pruebas de electroforesis, surgiendo la agarosa en 1969 como un candidato adecuado debido a las características que posee.
¿Cuál es el fundamento de la electroforesis en gel de agarosa para ácidos nucleicos?
La electroforesis es una técnica para la separación de biomoléculas (generalmente ácidos nucleicos o proteínas) en un campo eléctrico sobre una matriz porosa, cuya composición depende de la biomolécula a analizar.
Las moléculas en un campo eléctrico se mueven a una velocidad dependiente de su carga, forma y tamaño.
Las moléculas de ADN de las células son excesivamente grandes para avanzar a través de un gel de electroforesis normal, pero pueden analizarse si previamente se han fragmentado de forma controlada. Una de las formas más comunes de fragmentar el ADN es el empleo de endonucleasas de restricción.
La carga eléctrica de una molécula de DNA o RNA depende de sus grupos fosfato, y el número de éstos es igual al doble del número de pares de bases (pb). La forma de todas las moléculas de DNA es esencialmente la misma.
En consecuencia, resulta que la movilidad de los ácidos nucleicos en electroforesis depende exclusivamente de la longitud de la molécula.
¿Cuáles son los usos de la electroforesis en geles de agarosa en el ámbito de la biología molecular?
La electroforesis en geles de agarosa es el método estándar para:
- Separación de los fragmentos de ADN después de su digestión con enzimas de restricción.
- Separación de moléculas de ácidos nucleicos (ADN/ARN) antes de su transferencia a membranas para análisis posteriores.
- Análisis de los productos de PCR (reacción en cadena de la polimerasa) verificando si ocurrió o no amplificación.
- Estimación del tamaño de las moléculas en una mezcla de ADN o ARN.
- Estimación de la cantidad y/o la calidad de los ácidos nucleicos purificados.
¿Cuáles son las ventajas del uso de las Tabletas MaxAgarose?
Las tabletas MaxAgarose están diseñadas para proporcionar un entorno más limpio, seguro, organizado y conveniente que la agarosa en polvo. Cada tableta contiene una cantidad predeterminada de agarosa (0,5 g).
El uso de las tabletas MaxAgarose facilita la preparación de los geles para la electroforesis ya que no es necesario pesar la agarosa en polvo lo que requiere de una balanza y consume tiempo. Así mismo, evita derrames y la posibilidad de errores en la pesada.
Simplemente agregue la cantidad adecuada de tabletas a su tampón (buffer), incube a temperatura ambiente durante cinco minutos, caliente la solución y luego prepare su gel como de costumbre.
REFERENCIAS
- Electroforesis de proteínas y de ácidos nucleicos
Biomodel. Páginas de bioquímica y biología molecular (2021)
Universidad de Alcalá UHA. España.
http://biomodel.uah.es/tecnicas/elfo/inicio.htm
- Electroforesis: fundamento, técnica, para qué sirve, ejemplos.
Raquel Parada Puig. (29 de mayo de 2020).
Lifeder.com (life daily education & research).
https://www.lifeder.com/electroforesis/
- Electroforesis: fundamentos, avances y aplicaciones
Montalvo Navarro Carlos Antonio, Lugo Flores Marco Antonio
NISON / EPISTEMUS 26 / Año 13/ 2019/ pág.: 48-54
https://epistemus.unison.mx/index.php/epistemus/article/view/96/70
- Electroforesis de ADN
Francisco Fierro Fierro (2014)
Departamento de Biotecnología, División de Ciencias Biológicas y de la Salud, Universidad Autónoma Metropolitana-Unidad Iztapalapa, México D.F.
http://www2.inecc.gob.mx/publicaciones2/libros/710/electroforesis.pdf
- Separación de ADN por electroforesis horizontal en geles de agarosa
Manual de Prácticas de Laboratorio de Biología Molecular 2012
Universidad Autónoma Metropolitana
División de Ciencias Naturales e Ingeniería
http://www.cua.uam.mx/pdfs/conoce/libroselec/Manual_de_Prcticas_de_BM.pdf
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