Extensive restriction fragment length polymorphisms in a new isolate ofChlamydomonas reinhardtii

Gross, Christian H.; Ranum, Laura P.W.; Lefebvre, Paul

doi:10.1007/bf02427756

Cited by 98 publications

(69 citation statements)

References 20 publications

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“…Mapping the retrograde mutant strains: Each of the temperature-sensitive mutations in Chlamydomonas with decreased retrograde velocities and reduced numbers of retrograde particles was crossed to the polymorphic strain, CC-1952(Gross et al 1988 to map the mutant phenotypes with respect to a marker for each of the complex A subunits (IFT144, IFT140, IFT139, IFT122, IFT121, and IFT43), which were developed using sequences kindly provided by D. Cole and H. Qin (personal communication) (Table 2). In addition, if the gene was placed on a chromosome in the JGI assemblies (Merchant et al 2007), the position was verified by mapping to known markers.…”

Section: Resultsmentioning

confidence: 99%

Retrograde Intraflagellar Transport Mutants Identify Complex A Proteins With Multiple Genetic Interactions inChlamydomonas reinhardtii

Iomini¹,

et al. 2009

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The intraflagellar transport machinery is required for the assembly of cilia. It has been investigated by biochemical, genetic, and computational methods that have identified at least 21 proteins that assemble into two subcomplexes. It has been hypothesized that complex A is required for retrograde transport. Temperature-sensitive mutations in FLA15 and FLA17 show defects in retrograde intraflagellar transport (IFT) in Chlamydomonas. We show that IFT144 and IFT139, two complex A proteins, are encoded by FLA15 and FLA17, respectively. The fla15 allele is a missense mutation in a conserved cysteine and the fla17 allele is an in-frame deletion of three exons. The flagellar assembly defect of each mutant is rescued by the respective transgenes. In fla15 and fla17 mutants, bulges form in the distal one-third of the flagella at the permissive temperature and this phenotype is also rescued by the transgenes. These bulges contain the complex B component IFT74/72, but not a-tubulin or p28, a component of an inner dynein arm, which suggests specificity with respect to the proteins that accumulate in these bulges. IFT144 and IFT139 are likely to interact with each other and other proteins on the basis of three distinct genetic tests: (1) Double mutants display synthetic flagellar assembly defects at the permissive temperature, (2) heterozygous diploid strains exhibit second-site noncomplemention, and (3) transgenes confer two-copy suppression. Since these tests show different levels of phenotypic sensitivity, we propose they illustrate different gradations of gene interaction between complex A proteins themselves and with a complex B protein (IFT172).

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Section: Resultsmentioning

confidence: 99%

Retrograde Intraflagellar Transport Mutants Identify Complex A Proteins With Multiple Genetic Interactions inChlamydomonas reinhardtii

Iomini¹,

et al. 2009

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“…Many of these strains are ones that were contributed by members of the Chlamydomonas research community as their most frequently used wild-type strains (see Supplemental Data Set 1 for detailed strain histories). In addition to the 39 standard laboratory strains, one unrelated, but interfertile, isolate, CC-2290 (also known as S1 D2), was included for comparison (Gross et al, 1988). DNA was collected from the strains and sequenced on the Illumina platform.…”

Section: Genome Resequencing Reveals Genetic Diversity Of Laboratory mentioning

confidence: 99%

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

et al. 2015

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Chlamydomonas reinhardtii is a widely used reference organism in studies of photosynthesis, cilia, and biofuels. Most research in this field uses a few dozen standard laboratory strains that are reported to share a common ancestry, but exhibit substantial phenotypic differences. In order to facilitate ongoing Chlamydomonas research and explain the phenotypic variation, we mapped the genetic diversity within these strains using whole-genome resequencing. We identified 524,640 single nucleotide variants and 4812 structural variants among 39 commonly used laboratory strains. Nearly all (98.2%) of the total observed genetic diversity was attributable to the presence of two, previously unrecognized, alternate haplotypes that are distributed in a mosaic pattern among the extant laboratory strains. We propose that these two haplotypes are the remnants of an ancestral cross between two strains with ;2% relative divergence. These haplotype patterns create a fingerprint for each strain that facilitates the positive identification of that strain and reveals its relatedness to other strains. The presence of these alternate haplotype regions affects phenotype scoring and gene expression measurements. Here, we present a rich set of genetic differences as a community resource to allow researchers to more accurately conduct and interpret their experiments with Chlamydomonas.

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“…Mutations generated in this species can be mapped by crossing to the interfertile strain known as Chlamydomonas grossii, S1-D2 or its culture collection designation of CC-2290, which has a suitable profusion of sequence tagged sites (STS), cleavable amplified polymorphic sequence (CAPS), single nucleotide polymorphism (SNP), and RFLP markers (Gross et al, 1988;Vysotskaia et al, 2001;Grossman et al, 2003). Beginning with laborious RFLP-based mapping (Gross et al, 1988), Chlamydomonas mapping has moved toward a PCR-based method (Kathir et al, 2003) and now is poised to incorporate more high-throughput methods. This, in concert with an increasingly complete nuclear genome sequence (Grossman et al, 2003), provides the necessary tools for studies of all classes of mutations.…”

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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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Extensive restriction fragment length polymorphisms in a new isolate ofChlamydomonas reinhardtii

Cited by 98 publications

References 20 publications

Retrograde Intraflagellar Transport Mutants Identify Complex A Proteins With Multiple Genetic Interactions inChlamydomonas reinhardtii

Retrograde Intraflagellar Transport Mutants Identify Complex A Proteins With Multiple Genetic Interactions inChlamydomonas reinhardtii

Chlamydomonas Genome Resource for Laboratory Strains Reveals a Mosaic of Sequence Variation, Identifies True Strain Histories, and Enables Strain-Specific Studies

Beyond Complementation. Map-Based Cloning in Chlamydomonas reinhardtii

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