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Random sequences were generated by a fifth-order Markov chain based on 6-mer frequencies within the yeast genome.
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r). A polymerase chain reaction (PCR)-based HIS3 insertion scheme was used as described by A. Baudin, O. Ozier-Kalogeropoulos, A. Denouel, F. Lacroute, and C. Cullin [Nucleic Acids Res. 21, 3329 (1993)], with a protocol provided by L. Riles. Transformants growing on yeast extract, peptone, and dextrose His plates were picked and assayed by PCR for correct integration of the HIS3 marker gene replacing the target snoRNA.
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All computer codes, snoRNA search results, primer sequences, gel images, and other referenced data can be accessed on the WWW at rna.wustl.edu/snoRNAdb/
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The S. cerevisiae database is available at: genome-www.stanford.edu/Saccharomyces/
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0345281790
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note
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We thank L. Lutfiyya and L. Riles from the M. Johnston lab for protocols and guidance in all aspects of yeast handling, gene disruptions, and colony PCR and RNA preparations; M. Fournier, J. Ni, D. Samarsky, and B. E. H. Maden for helpful discussions and sharing of unpublished observations; S. Johnson and L. Lutfiyya for careful reading of the manuscript; and M. Johnston for providing the haploid strain yM4585 and diploid strain yM4587. Supported by NIH grant R01-HG01363 and by a gift from Eli Lilly.
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