Heterogeneity in Bacillus cereus PCR products detected by ESI-FTICR mass spectrometry.
PCR amplification of a segment of the 16/23S rDNA interspace region (ISR) from Bacillus cereus 6464 produced a mixture of products. An 89-bp product was predicted on the basis of the reported sequence. The ESI-FTICR analysis revealed three double-stranded products, differing in size by a single nucleotide corresponding to two homoduplexes of 89 and 88 base pairs and a heteroduplex of 89 and 88 nucleotide strands. These were produced from a single preparation of genomic DNA and a single primer pair. ESI-FTICR analysis of the single strands identified a deletion of a T in the coding strand and a corresponding loss of an A in the noncoding strand of this product. The ESI-FTICR analysis indicated the presence of an unreported sequence variation between rRNA operons in this organism. This report illustrates that PCR products amplified from templates differing by a single nucleotide can be resolved and identified using ESI-FTICR at the 89-bp level. Furthermore, the ESI-FTICR mass measurements provided the identity of the deletion, which is indicative of interoperon variability.
Wunschel
D S
DS
Macromolecular Structure and Dynamics, Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington 99352, USA.
Muddiman
D C
DC
Fox
K F
KF
Fox
A
A
Smith
R D
RD
D001409:Bacillus cereus / Q000235:genetics*; D017357:Cyclotrons; D004269:DNA, Bacterial / Q000032:analysis*; D005583:Fourier Analysis; D013058:Mass Spectrometry / Q000379:methods*; D016133:Polymerase Chain Reaction
D004269:DNA, Bacterial