Determination of microbial genome sizes by two-dimensional denaturing gradient gel electrophoresis.
In two-dimensional denaturing gradient gel electrophoresis, DNA is digested with a restriction endonuclease and the resulting DNA fragments are separated as a function of size by conventional agarose gel electrophoresis. Following this first dimension electrophoresis, the fragment distribution is placed at the top of a denaturing gradient slab gel and electrophoresis is carried out parallel to the gradient direction. This second dimension separation is a complex function of the base sequence of each fragment. Analysis of the DNA fragment distribution as a function of fragment size allows the DNA size to be calculated. This method has been applied to calculate three microbial genome sizes: Mycoplasma capricolum, 724 kb; Acholeplasma laidlawii, 1646 kb; and Hemophilus influenzae, 1833 kb.
Poddar
S K
SK
Department of Microbiology and Immunology, University of Rochester, NY 14642.
Maniloff
J
J
D000127:Acholeplasma / Q000235:genetics; D001482:Base Composition; D004269:DNA, Bacterial / Q000032:analysis*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006193:Haemophilus influenzae / Q000235:genetics; D008970:Molecular Weight; D009174:Mycoplasma / Q000235:genetics; D009691:Nucleic Acid Denaturation
D004269:DNA, Bacterial