CHAPTER 8 



The Electrophoretic Separation of Nucleic Acid Components 



J. D. SMITH 



Page 

 I. Theory of the Electrophoretic Separation of Nucleic Acid Components. 267 



1. General Considerations 267 



2. Separation of Nucleotides and Polynucleotides 268 



3. Purines, Pyrimidines, and Nucleosides 272 



II. Apparatus and Techniques 274 



III. Applications of the Method 276 



1. Determination of Nucleic Acid Composition 276 



2. Separation of Nucleotides from Other Phosphorus Compounds .... 277 



3. Isolation of Nucleotides and Polynucleotides from Enzymic and Partial 

 Chemical Hydrolysates of Nucleic Acids 279 



a. Cyclic Ribonucleotides 279 



b. Polynucleotides from Ribonucleic Acid 280 



c. Polynucleotides and Dinucleoside Monophosphates from Deoxyribo- 

 nucleic Acid 282 



d. Nucleoside Diphosphates 282 



4. Separation of Nucleosides and Nucleotides as Borate Complexes . . 283 



Nucleic acids and their components bear a number and variety of ioniz- 

 able groups; and it is natural that electrophoretic techniques should play 

 an important part in their separation. The development of electrophoresis 

 on paper has greatly facilitated separations of nucleic acid derivatives which 

 would be impossible by paper chromatography and time-consuming by 

 ion-exchange methods. All the manipulations can be carried out on very 

 small amounts of material. An important feature of the method is that the 

 relative mobilities of nucleic acid components at any given pH may be 

 predicted quite accurately, thus aiding in the identification of unknown 

 components. 



I. Theory of the Electrophoretic Separation of Nucleic Acid Components 



1. Gener.\l Considerations 



A molecule in a fluid subjected to a voltage gradient (E) is acted upon 

 by a force equal to (EQ), where Q is the net charge on the molecule and is 

 given by the algebraic sum of the products of the number of ionizing groups 



267 



