High-density microarray of small-subunit ribosomal DNA probes.
Ribosomal DNA sequence analysis, originally conceived as a way to provide a universal phylogeny for life forms, has proven useful in many areas of biological research. Some of the most promising applications of this approach are presently limited by the rate at which sequences can be analyzed. As a step toward overcoming this limitation, we have investigated the use of photolithography chip technology to perform sequence analyses on amplified small-subunit rRNA genes. The GeneChip (Affymetrix Corporation) contained 31,179 20-mer oligonucleotides that were complementary to a subalignment of sequences in the Ribosomal Database Project (RDP) (B. L. Maidak et al., Nucleic Acids Res. 29:173-174, 2001). The chip and standard Affymetrix software were able to correctly match small-subunit ribosomal DNA amplicons with the corresponding sequences in the RDP database for 15 of 17 bacterial species grown in pure culture. When bacteria collected from an air sample were tested, the method compared favorably with cloning and sequencing amplicons in determining the presence of phylogenetic groups. However, the method could not resolve the individual sequences comprising a complex mixed sample. Given these results and the potential for future enhancement of this technology, it may become widely useful.
Wilson
Kenneth H
KH
Veterans Affairs Medical Center and Duke University Medical Center, Durham, North Carolina 27710, USA.
Wilson
Wendy J
WJ
Radosevich
Jennifer L
JL
DeSantis
Todd Z
TZ
Viswanathan
Vijay S
VS
Kuczmarski
Thomas A
TA
Andersen
Gary L
GL
D000391:Air Microbiology; D016360:Clostridioides difficile / Q000235:genetics; D017124:Conserved Sequence; D004275:DNA, Ribosomal / Q000032:analysis*; D030561:Databases, Nucleic Acid; D016952:Legionella pneumophila / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D010802:Phylogeny; D013211:Staphylococcus aureus / Q000235:genetics
D004275:DNA, Ribosomal