Molecular markers for rapidly identifying candidate genes in Chlamydomonas reinhardtii. Ery1 and ery2 encode chloroplast ribosomal proteins.
To take advantage of available expressed sequence tags and genomic sequence, we have developed 64 PCR-based molecular markers in Chlamydomonas reinhardtii that map to the 17 linkage groups. These markers will allow the rapid association of a candidate gene sequence with previously identified mutations. As proof of principle, we have identified the genes encoded by the ERY1 and ERY2 loci. Mendelian mutations that confer resistance to erythromycin define three unlinked nuclear loci in C. reinhardtii. Candidate genes ribosomal protein L4 (RPL4) and L22 (RPL22) are tightly linked to the ERY1 locus and ERY2 locus, respectively. Genomic DNA for RPL4 from wild type and five mutant ery1 alleles was amplified and sequenced and three different point mutations were found. Two different glycine residues (G(102) and G(112)) are replaced by aspartic acid and both are in the unstructured region of RPL4 that lines the peptide exit tunnel of the chloroplast ribosome. The other two alleles change a splice site acceptor site. Genomic DNA for RPL22 from wild type and three mutant ery2 alleles was amplified and sequenced and revealed three different point mutations. Two alleles have premature stop codons and one allele changes a splice site acceptor site.
Bowers
Amber K
AK
Department of Genetics, Washington University School of Medicine, 660 S. Euclid Avenue, St Louis, MO 63110, USA.
Keller
Jennifer A
JA
Dutcher
Susan K
SK
D000483:Alleles; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism; D015415:Biomarkers; D016825:Chlamydomonas reinhardtii / Q000187:drug effects / Q000235:genetics*; D002736:Chloroplasts / Q000737:chemistry / Q000235:genetics*; D018388:Codon, Terminator; D004275:DNA, Ribosomal / Q000235:genetics; D004352:Drug Resistance, Microbial / Q000235:genetics; D004917:Erythromycin / Q000494:pharmacology; D005796:Genes; D008040:Genetic Linkage; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017354:Point Mutation; D022821:RNA Splice Sites; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics*; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D015415:Biomarkers; D018388:Codon, Terminator; D004275:DNA, Ribosomal; D022821:RNA Splice Sites; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; C056368:ribosomal protein L4; D001224:Aspartic Acid; D004917:Erythromycin