Differentiation of G:C vs A:T and G:C vs G:mC Base Pairs in the Latch Zone of α-Hemolysin.
The α-hemolysin (α-HL) nanopore can detect DNA strands under an electrophoretic force via many regions of the channel. Our laboratories previously demonstrated that trapping duplex DNA in the vestibule of wild-type α-HL under force could distinguish the presence of an abasic site compared to a G:C base pair positioned in the latch zone at the top of the vestibule. Herein, a series of duplexes were probed in the latch zone to establish if this region can detect more subtle features of base pairs beyond the complete absence of a base. The results of these studies demonstrate that the most sensitive region of the latch can readily discriminate duplexes in which one G:C base pair is replaced by an A:T. Additional experiments determined that while neither 8-oxo-7,8-dihydroguanine nor 7-deazaguanine opposite C could be differentiated from a G:C base pair, in contrast, the epigenetic marker 5-methylcytosine, when present in both strands of the duplex, yielded new blocking currents when compared to strands with unmodified cytosine. The results are discussed with respect to experimental design for utilization of the latch zone of α-HL to probe specific regions of genomic samples.
Department of Chemistry, University of Utah , 315 South 1400 East, Salt Lake City, Utah 84112-0850, United States.
Yun
Y
Ding
Department of Chemistry, University of Utah , 315 South 1400 East, Salt Lake City, Utah 84112-0850, United States.
Aaron M
AM
Fleming
Department of Chemistry, University of Utah , 315 South 1400 East, Salt Lake City, Utah 84112-0850, United States.
Henry S
HS
White
Department of Chemistry, University of Utah , 315 South 1400 East, Salt Lake City, Utah 84112-0850, United States.
Cynthia J
CJ
Burrows
D020029:Base Pairing; D004247:DNA / Q000235:genetics; D004560:Electricity; D006460:Hemolysin Proteins / Q000737:chemistry*; D058608:Nanopores
D006460:Hemolysin Proteins; D004247:DNA
base pair detection; duplex DNA; epigenetic marker detection; latch zone; α-hemolysin