Characterization of a circular plasmid from the yeast Kluyveromyces waltii

Chen, Xin J.; Cong, Yu Sheng; Wésolowski-Louvel, Micheline; Li, Yu Y.; Fukuhara, Hiroshi

doi:10.1099/00221287-138-2-337

Cited by 22 publications

(15 citation statements)

References 19 publications

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“…Only reads mapping uniquely in the L. waltii genome were analyzed. The L. waltii genome used here includes the assembled L. waltii genome described previously (Di Rienzi et al 2011), the L. waltii mating locus, the 2 mm plasmid (Chen et al 1992), and the rDNA locus (described here; see Supplemental Text). For aligning paired-end reads, an insert size between 250 and 1100 was allowed in Bowtie.…”

Section: Illumina Sequence Data Analysismentioning

confidence: 99%

Maintaining replication origins in the face of genomic change

et al. 2012

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Origins of replication present a paradox to evolutionary biologists. As a collection, they are absolutely essential genomic features, but individually are highly redundant and nonessential. It is therefore difficult to predict to what extent and in what regard origins are conserved over evolutionary time. Here, through a comparative genomic analysis of replication origins and chromosomal replication patterns in the budding yeasts Saccharomyces cerevisiae and Lachancea waltii, we assess to what extent replication origins survived genomic change produced from 150 million years of evolution. We find that L . waltii origins exhibit a core consensus sequence and nucleosome occupancy pattern highly similar to those of S. cerevisiae origins. We further observe that the overall progression of chromosomal replication is similar between L. waltii and S. cerevisiae. Nevertheless, few origins show evidence of being conserved in location between the two species. Among the conserved origins are those surrounding centromeres and adjacent to histone genes, suggesting that proximity to an origin may be important for their regulation. We conclude that, over evolutionary time, origins maintain sequence, structure, and regulation, but are continually being created and destroyed, with the result that their locations are generally not conserved.

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Section: Illumina Sequence Data Analysismentioning

confidence: 99%

Maintaining replication origins in the face of genomic change

et al. 2012

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“…[9]. Our results show that the Kw protein is alone sufficient to promote intermolecular and intramolecular recombination in vitro of a DNA target site consisting of two 18-bp elements separted by a 7-bp spacer.…”

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confidence: 68%

“…The 2 p-like plasmid, pKWl from K. waltii contains four ORF A, B, C and D. Chen et al [9] suggested that the A gene encodes a recombinase, on the basis of similarity of its deduced amino acid sequence with that of the FLP gene of 2 p and its equivalent in other yeast plasmids. Like other 2 p plasmids, pKWl has two inverted repeat sequences of about 300 bp.…”

Section: Resultsmentioning

confidence: 99%

“…Like other 2 p plasmids, pKWl has two inverted repeat sequences of about 300 bp. Within this region, the putative recombination target sites have been identified by Chen et al [9] on the basis of their sequence structure. Each putative target site contains three 18-bp perfect repeat elements, two of which are inverted, separated by a non-palindromic stretch of 7-bp.…”

Section: Resultsmentioning

confidence: 99%

“…pBSKw contains the coding sequence of the Kw recombinase isolated as a BspEI-Scal partial fragment from plasmid pKWS1 [9] ligated into pBluescribe (Stratagene) downstream of the T3 promoter.…”

Section: Methodsmentioning

confidence: 99%

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The Kw Recombinase, an Integrase from Kluyveromyces Waltii

Ringrose

Angrand

Stewart

1997

European Journal of Biochemistry

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Site-specific recombinases of the integrase family share limited amino-acid-sequence similarity, but use a common reaction mechanism to recombine distinct DNA target sites. Here we report the characterisation of the Kw site-specific recombinase, encoded on the 2 p-like plasmid pKWS1 from the yeast Kluyveromyces waltii. Using in vitro-translated Kw recombinase, we show that the protein is able to bind and to recombine its putative DNA target site. Recombination is conservative and the Kw target site has a spacer of seven base pairs. We show that Kw recombinase is able to mediate recombination in a mammalian cell line, thus, it has potential for use as a tool for genomic manipulation in heterologous systems.Keywords: site-specific recombinase ; FLP recombinase ; Kluyveromyces waltii, Kw recombinase.Site-specific recombinases mediate DNA rearrangements at specific target sites, and can be classified into several distinct families on the basis of their amino acid sequences and certain features of their reaction mechanisms. The integrase family of site-specific recombinases shows limited amino-acid-sequence similarity ([ 1, 21 ; Fig. I), but those members that have been characterised share four catalytic residues and use a common reaction mechanism to recombine distinct DNA target sites in various biological contexts. Well-characterised examples of this family are i integrase from bacteriophage L, and Cre recombinase from bacteriophage PI 13). A further subfamily within the integrase family is the yeast family, whose members share somewhat higher amino-acid-sequence similarity [4]. The best characterised member of this family is the FLP site-specific recombinase encoded on the 2 p circle from Succharomyces cerevisiae [5]. A second member, the R recombinase from Zygosaccharomyces rouxii, has been characterised in vivo [6] and in vitro [7]. By analogy to the FLP system, other yeast-family recombinases and their recombination target sites have been identified from the 2 p-like plasmids of other yeast species [4, 8, 91. Common features that have emerged from the characterisation of the FLP and R recombinases and their target sites are that the recombination reaction is carried out by a single protein, and that the minimal target site required for efficient recombination is quite simple, consisting of two 12-bp or 13-bp repeats in inverted orientation, separated by a non-palindromic spacer of 7 bp or 8 bp [6,7,10, 111. The outcome of recombination between two DNA target sites is dictated by the orientation of the spacers with respect to one another 1121. Furthermore, these recombinases catalyse intramolecular recombination (excision or inversion) and intermolecular recombination (integration or translocation). All of the above features are shared by the Crerecombinase system from bacteriophage P1. These characteristics have proved to be useful in the development of genomic- engineering strategies, in which all three recombinases have been used to direct DNA rearrangements in living organisms, including bacteria, yeast, ...

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Genetic, genomic, and molecular tools for studying the protoploid yeast, L. waltii

Rienzi

Lindstrom

Lancaster

et al. 2010

Yeast

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Sequencing of the yeast Kluyveromyces waltii (recently renamed Lachancea waltii ) provided evidence of a whole genome duplication event in the lineage leading to the well-studied Saccharomyces cerevisiae. While comparative genomic analyses of these yeasts have proven to be extremely instructive in modeling the loss or maintenance of gene duplicates, experimental tests of the ramifications following such genome alterations remain difficult. To transform L. waltii from an organism of the computational comparative genomic literature into an organism of the functional comparative genomic literature, we have developed genetic, molecular and genomic tools for working with L. waltii. In particular, we have characterized basic properties of L. waltii (growth, ploidy, molecular karyotype, mating type and the sexual cycle), developed transformation, cell cycle arrest and synchronization protocols, and have created centromeric and non-centromeric vectors as well as a genome browser for L. waltii. We hope that these tools will be used by the community to follow up on the ideas generated by sequence data and lead to a greater understanding of eukaryotic biology and genome evolution.

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Characterization of a circular plasmid from the yeast Kluyveromyces waltii

Cited by 22 publications

References 19 publications

Maintaining replication origins in the face of genomic change

Maintaining replication origins in the face of genomic change

The Kw Recombinase, an Integrase from Kluyveromyces Waltii

Genetic, genomic, and molecular tools for studying the protoploid yeast, L. waltii

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