Characterization of synthetic DNA bar codes in Saccharomyces cerevisiae gene-deletion strains.
Incorporation of strain-specific synthetic DNA tags into yeast Saccharomyces cerevisiae gene-deletion strains has enabled identification of gene functions by massively parallel growth rate analysis. However, it is important to confirm the sequences of these tags, because mutations introduced during construction could lead to significant errors in hybridization performance. To validate this experimental system, we sequenced 11,812 synthetic 20-mer molecular bar codes and adjacent sequences (>1.8 megabases synthetic DNA) by pyrosequencing and Sanger methods. At least 31% of the genome-integrated 20-mer tags contain differences from those originally synthesized. However, these mutations result in anomalous hybridization in only a small subset of strains, and the sequence information enables redesign of hybridization probes for arrays. The robust performance of the yeast gene-deletion dual oligonucleotide bar-code design in array hybridization validates the use of molecular bar codes in living cells for tracking their growth phenotype.
Eason
Robert G
RG
Stanford Genome Technology Center, 855 California Avenue, Palo Alto, CA 94304, USA.
Pourmand
Nader
N
Tongprasit
Waraporn
W
Herman
Zelek S
ZS
Anthony
Kevin
K
Jejelowo
Olufisayo
O
Davis
Ronald W
RW
Stolc
Viktor
V
D017931:DNA Primers; D004271:DNA, Fungal / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics*; D017353:Gene Deletion; D005800:Genes, Fungal / Q000235:genetics; D009154:Mutation; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D017931:DNA Primers; D004271:DNA, Fungal