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AAT Bioquest

What are the methods used to transfer separated DNA from gels to solid supports?

Posted May 2, 2024


Answer

Electrophoretic transfer (also known as electroblotting) is a method used to transfer molecules such as DNA or proteins from a gel matrix onto a solid support membrane. There are two main variations of electrophoretic transfer: semi-dry transfer and wet transfer. In semi-dry transfer, the gel and membrane are sandwiched between layers of buffer-saturated materials, such as filter paper or sponges, and then subjected to an electric field. This method typically utilizes less buffers and requires shorter transfer times compared to wet transfer. 

In wet transfer, the gel and membrane are submerged in a buffer solution within a transfer apparatus, such as a tank or cassette, and an electric current is applied. This method allows for more uniform transfer of molecules and is generally preferred for larger proteins or DNA fragments. Wet transfer is considered more reliable as it minimizes the risk of the gel drying out during transfer, which can lead to incomplete or uneven transfer.

In semi-dry transfer, the gel and membrane sandwich is placed horizontally between two plate electrodes to improve transfer speed. By limiting the amount of buffer to what's in the sandwich, the current is maximized through the gel and not around it. 

In diffusion blotting, molecules are transferred from a gel matrix to a transfer membrane due to their natural movement from areas of high concentration to low concentration. During blotting, proteins diffuse out of the gel and are absorbed onto the membrane. 

Additional resources

Western Blot: Technique, Theory, and Trouble Shooting

Western Blotting Assays

Gel Electrophoresis

Gelite™ Green Nucleic Acid Gel Staining Kit