BioCoder #4

Beer Bottle Minipreps

Joe Rokicki

Introduction

Minipreps are the bread and butter of the genetic engineer. After constructing or procuring a DNA sequence of interest, one of the first next steps is often to insert that DNA into a larger circular piece of DNA called a plasmid. We do this because plasmids have the wonderful property of being autonomously amplified inside of bacteria. If you can get your DNA of interest into a plasmid and you can get that plasmid into a bacterium, all of the fancy, error-correcting, DNA copying machinery of the bacterium will go to work for you, automatically copying your DNA. You can sit back and relax and grow up a bacterial broth that will be rich in your DNA sequence. The only thing you have to worry about is breaking the cells open and purifying your amplified DNA back out, away from all the other genomic DNA, RNA, proteins, and miscellaneous cellular debris. This is where the miniprep comes in.

A miniprep is a column-based plasmid purification protocol where crude cell lysate is passed through a resin that specifically binds plasmid DNA and lets everything else pass through. The plasmid DNA is later released from the column in a highly pure and concentrated form. The miniprep’s speed, ease of use, and purity have made it the de facto standard over other plasmid purification protocols such as phenol chloroform extractions and cesium chloride gradients. Yet, despite being one of the most universally performed protocols in molecular biology, the miniprep ...