Analysis of <i>Chlamydomonas reinhardtii</i> Genome Structure Using Large‐Scale Sequencing of Regions on Linkage Groups I and III

Li, Jin Billy; Lin, Shaoping; Jia, Honggui; Wu, Hongmin; Roe, Bruce A.; Kulp, David; Stormo, Gary D.; Dutcher, Susan K.

doi:10.1111/j.1550-7408.2003.tb00109.x

Cited by 25 publications

(16 citation statements)

References 49 publications

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“…C. reinhardtii has a compact mitochondrial genome containing no introns and 10% intergenic DNA, not counting the terminal repeat sequences (Gray and Boer 1988;Michaelis et al 1990), while the proportion of intron plus intergenic DNA in the nuclear genome is 70% on the basis of linkage group III sequence (Li et al 2003). This dissimilarity in genome compactness cannot be explained by mutation rate differences because the estimated mutation rates in the two genetic compartments seem to be quite similar.…”

Section: Trpmentioning

confidence: 80%

Mitochondrial Genome Sequence Evolution in Chlamydomonas

Popescu

Lee

2007

Genetics

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The mitochondrial genomes of the Chlorophyta exhibit significant diversity with respect to gene content and genome compactness; however, quantitative data on the rates of nucleotide substitution in mitochondrial DNA, which might help explain the origin of this diversity, are lacking. To gain insight into the evolutionary forces responsible for mitochondrial genome diversification, we sequenced to near completion the mitochondrial genome of the chlorophyte Chlamydomonas incerta, estimated the evolutionary divergence between Chlamydomonas reinhardtii and C. incerta mitochondrial protein-coding genes and rRNA-coding regions, and compared the relative evolutionary rates in mitochondrial and nuclear genes. Synonymous and nonsynonymous substitution rates do not differ significantly between the mitochondrial and nuclear protein-coding genes. The mitochondrial rRNA-coding regions, however, are evolving much faster than their nuclear counterparts, and this difference might be explained by relaxed functional constraints on the mitochondrial translational apparatus due to the small number of proteins synthesized in Chlamydomonas mitochondria. Substitution rates at synonymous sites in a nonstandard mitochondrial gene (rtl) and at intronic and synonymous sites in nuclear genes expressed at low levels suggest that the mutation rate is similar in these two genetic compartments. Potential evolutionary forces shaping mitochondrial genome evolution in Chlamydomonas are discussed.

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Section: Trpmentioning

confidence: 80%

Mitochondrial Genome Sequence Evolution in Chlamydomonas

Popescu

Lee

2007

Genetics

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“…In addition, the gene prediction programs used have a significant error frequency, both in defining splice sites and in the recognition of mitochondrial and chloroplast transit peptides. Annotation has been aided by the combined EST and protein homolog data displayed on the genome browser, but only about 60% of the predicted genes have EST coverage and 42% have protein homologies (Li et al, 2003). These considerations, of course, are not unique to Chlamydomonas.…”

Section: Future Perspectivesmentioning

confidence: 99%

Beyond Complementation. Map-Based Cloning in Chlamydomonas reinhardtii

et al. 2005

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Chlamydomonas reinhardtii is an excellent model system for plant biologists because of its ease of manipulation, facile genetics, and the ability to transform the nuclear, chloroplast, and mitochondrial genomes. Numerous forward genetics studies have been performed in Chlamydomonas, in many cases to elucidate the regulation of photosynthesis. One of the resultant challenges is moving from mutant phenotype to the gene mutation causing that phenotype. To date, complementation has been the primary method for gene cloning, but this is impractical in several situations, for example, when the complemented strain cannot be readily selected or in the case of recessive suppressors that restore photosynthesis. New tools, including a molecular map consisting of 506 markers and an 8X-draft nuclear genome sequence, are now available, making map-based cloning increasingly feasible. Here we discuss advances in map-based cloning developed using the strains mcd4 and mcd5, which carry recessive nuclear suppressors restoring photosynthesis to chloroplast mutants. Tools that have not been previously applied to Chlamydomonas, such as bulked segregant analysis and marker duplexing, are being implemented to increase the speed at which one can go from mutant phenotype to gene. In addition to assessing and applying current resources, we outline anticipated future developments in map-based cloning in the context of the newly extended Chlamydomonas genome initiative.Sometimes called green yeast (Goodenough, 1992), the unicellular, eukaryotic green alga Chlamydomonas reinhardtii (hereafter called Chlamydomonas) is a venerable model system for plant biology as well as for cell motility. The tag green yeast refers to its haploid vegetative state, the existence of two mating types, and the general similarity in applicable genetic techniques. These aspects of Chlamydomonas biology have been previously reviewed (Rochaix, 1995).Like many microorganisms, screening of Chlamydomonas strains for rare mutations is straightforward, since large numbers of cells can be plated on an appropriate selective medium, or nonswimmers, for example, can be selected from large numbers of swimming cells. At the same time, the ease of nuclear transformation in Chlamydomonas, coupled with the plant-like nonhomologous integration of transforming DNA, facilitates the creation of insertional mutant collections. Taken together, the assortment of techniques useable for Chlamydomonas indulges both the amateur and experienced geneticist, yielding sometimes overwhelming collections of mutant strains. In this report, we focus on mutants affecting photosynthesis, in keeping with the thrust of this journal, and the emphasis of the newly renewed and National Science Foundation-supported Chlamydomonas genome project (http://www.chlamy.org/). However, the map-based cloning tools described here are generally applicable to Chlamydomonas biology.The use of Chlamydomonas to study the elaboration and regulation of the photosynthetic apparatus is long established and was recently review...

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“…The COILS (www.ch.embnet.org/software/COILS_form.html) and PairCoil (http://paircoil.lcs.mit.edu/cgi-bin/paircoil) servers were used to predict coiled-coil regions in the Oda5 protein (Lupas et al, 1991;Berger et al, 1995). To predict regions of coding potential, the ODA5 and AK genomic sequences were analyzed using the GreenGenie gene prediction program (http:// www.cse.ucsc.edu/%7Edkulp/cgi-bin/greenGenie) (Li et al, 2003). The AK protein was examined using the Joint Genome Institute (Walnut Creek, CA; JGI) Chlamydomonas version 2.0 genome database (http://shake.jgi-psf.org/chlre2/ chlre2.home.html), and the PROSITE protein families and domains database (http://us.expasy.org/prosite/).…”

Section: Computational Analysismentioning

confidence: 99%

“…To identify potential coding regions, we analyzed the sequence using the GreenGenie gene prediction program (Li et al, 2003) (Figure 1A). PCR primers designed from the predicted exons were used to amplify the ODA5 cDNA from wild-type Chlamydomonas cDNA libraries.…”

Section: Cloning the Oda5 Cdnamentioning

confidence: 99%

Oda5p, a Novel Axonemal Protein Required for Assembly of the Outer Dynein Arm and an Associated Adenylate Kinase

Wirschell

Pazour

Yoda

et al. 2004

MBoC

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Of the uncloned ODA genes required for outer dynein arm assembly in Chlamydomonas, ODA5 and ODA10 are of particular interest because they do not encode known subunits of the outer arm or the outer dynein arm-docking complex (ODA-DC), and because genetic studies suggest their products interact. Beginning with a tagged oda5 allele, we isolated genomic and cDNA clones of the wild-type gene. ODA5 predicts a novel, 66-kDa coiled-coil protein. Immunoblotting indicates Oda5p is an axonemal component that assembles onto the axoneme independently of the outer arm and ODA-DC and is uniquely missing in oda5 and oda10 axonemes. Oda5p is released from the axoneme by extraction with 0.6 M KCl, but the soluble Oda5p does not cosediment with the outer dynein arm/ODA-DC in sucrose gradients. Quantitative mass spectrometry by using isotope coded affinity tagging revealed that a previously unidentified adenylate kinase is reduced 35-50% in oda5 flagella. Direct enzymatic assays demonstrated a comparable reduction in adenylate kinase activity in oda5 flagella, and also in oda10 flagella, but not in flagella of other oda mutants. We propose that Oda5p is part of a novel axonemal complex that is required for outer arm assembly and anchors adenylate kinase in proximity to the arm.

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Analysis of Chlamydomonas reinhardtii Genome Structure Using Large‐Scale Sequencing of Regions on Linkage Groups I and III

Cited by 25 publications

References 49 publications

Mitochondrial Genome Sequence Evolution in Chlamydomonas

Mitochondrial Genome Sequence Evolution in Chlamydomonas

Beyond Complementation. Map-Based Cloning in Chlamydomonas reinhardtii

Oda5p, a Novel Axonemal Protein Required for Assembly of the Outer Dynein Arm and an Associated Adenylate Kinase

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