Precast gels

The equipment and reagents for gel electrophoresis are readily available and familiar to laboratory workers. Particularly noteworthy is the steady increase in the popularity of precast polyacrylamide gels since their introduction in the early 1990s (see Section 8.2.4). Precast gels provide researchers with off-the-shelf convenience and reproducibility and help to make gel electrophoresis and IEF commonplace laboratory practices. 

Precast gel Real Gel Plate (Bio Craft, Tokyo, Japan) Store at 4°C. 

Precast Gels As previously mentioned, precast gels of polyacrylamide and agarose are now commercially available. A wide variety of gel sizes, types, configurations, and compositions may be purchased. Costs are... 

Reusable Precast Gels Precast agarose gels are now available that may be recycled. After a run, the DNA samples on the gel are removed by reversing the direction of the electric field. The gels are then reloaded with new samples and reused. 

M. Brush, The Scientist, July 20, 18-21 (1998). A Profile of Precast Acrylamide Gels. M. Brush, The Scientist, June 7, 15-17 (1999). A Profile of New Precast Gels for Nucleic Acid Analysis. ... 

It is recommended that precast gels be obtained commercially (Jule or Novex). They are cost- and time-effective and they come in a variety of sizes and types. If students are to make their own gels, the following items, reagents, and solutions should be provided. Many types of gel casters are commercially available. 

Glass plates, spacers, comb, electrical tape (2 inch diameter), and clamps or precast gels of 12% acrylamide 0.75 mm thickness, 8 X 10 cm. 

If precast gels are provided, move ahead to the preparation of protein samples for electrophoresis. If gels are to be made, follow these instructions for preparation of an SDS gel of 12% acrylamide. 

Mix components in a 0.5-mL Eppendorf tube and heat in a boiling water bath for 3 minutes to denature the proteins. Cool to room temperature and apply 10 pL of a sample to a well on the gel. Use gel-loading tips on an automatic pipettor to deliver these small samples. One or two wells should contain molecular weight standards. Be sure to record placement of each protein and standards in a sample well in your notebook. Follow the instructions provided with your electrophoresis apparatus and by the supplier of precast gels. 

It is recommended that students be provided precast gels for this exper iment, especially if they prepared their own in Experiment 4. Availability of preformed gels will ensure completion of the experiment in two 3-hour pc riods and will allow students to learn new procedures and techniques. 

Students may be instructed to prepare their own agarose gels or precast gels may be purchased. If precast gels are provided, part C requires approximately 1 hour. An additional hour is required if gels are made by students. 

This protocol can be used for either standard (14 x 16-cm) or mini-format (8 x 10-cm) gels. The recipes can be adjusted for 0.75 or 1.5-mm-thick gels. Alternatively, precast gels for SDS-PAGE, which are available from a number of manufacturers, can be used, and protocol should be started at step 16. 

---