Methods to electrophoretically stretch DNA: microcontractions, gels, and hybrid gel-microcontraction devices.
The ability to controllably and continuously stretch large DNA molecules in a microfluidic format is important for gene mapping technologies such as Direct Linear Analysis (DLA). We have recently shown that electric field gradients can be readily generated in a microfluidic device and the resulting field is purely elongational. We present a single molecule fluorescence microscopy analysis of T4 DNA (169 kbp) stretching in the electric field gradients in a hyperbolic contraction microchannel. In addition, we are able to selectively pattern a crosslinked gel anywhere inside the microchannel. With an applied electric field, DNA molecules are forced to reptate through the gel and they moderately stretch as they exit the gel. By placing a gel immediately in front of the hyperbolic contraction, we bypass "molecular individualism" and achieve highly uniform and complete stretching of T4 DNA.
Department of Chemical Engineering, Massachusetts Institute of Technology, MIT Room 66-456, 77 Massachusetts Ave., Cambridge, MA 02139, USA.
Greg C
GC
Randall
Kelly M
KM
Schultz
Patrick S
PS
Doyle
D017122:Bacteriophage T4 / Q000737:chemistry / Q000235:genetics; D004279:DNA, Viral / Q000737:chemistry*; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods*; D005782:Gels / Q000737:chemistry*; D008832:Microchemistry / Q000295:instrumentation / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D009690:Nucleic Acid Conformation
D004279:DNA, Viral; D005782:Gels