Enhancement of Somatic Intrachromosomal Homologous Recombination in Arabidopsis by the HO Endonuclease

Chiurazzi, Maurizio; Ray, Animesh; Viret, Jean-Frédéric; Perera, Ranjan J.; Wang, Xiaohui; Lloyd, A. C.; Signer, Ethan R.

doi:10.2307/3870412

Cited by 17 publications

(18 citation statements)

References 35 publications

(49 reference statements)

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“…This frequency was determined by relating the number of recombination events (sectors) to the number of genomes present per plant at the time of staining, as described by Swoboda et al (1994). This value is similar to that reported previously for the rate of spontaneous recombination of a GU.US transgene similar to the GU-Ds-US transgene used here, but which lacks a Ds insertion between the GU and US segments (Swoboda et al 1994;Chiurazzi et al 1996). In the presence of the sAc transposase source, the average recombination frequency of the GU-Ds-US construct is increased by more than 1000 fold.…”

Section: Detection Of Somatic Recombination Events In Transgenic Plantssupporting

confidence: 58%

“…Moreover, homologous recombination in plants is enhanced by in vivo induction of DNA double-strand breaks by a site-speci®c endonuclease (Puchta et al 1993;Puchta 1998). In Arabidopsis, generation of DSBs by HO endonuclease increased the frequency of somatic intrachromosomal homologous recombination about tenfold (Chiurazzi et al 1996). Therefore, transient DNA double strand breaks generated by transposon excision may be a critical initiator of recombination events.…”

Section: Discussionmentioning

confidence: 99%

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Intrachromosomal homologous recombination in Arabidopsis induced by a maize transposon

Xiao

Peterson

2000

Mol Gen Genet

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In plants, the frequency of spontaneous intrachromosomal homologous recombination is low. Here, we show that a maize transposable element greatly stimulates intrachromosomal homologous recombination between direct repeat sequences in Arabidopsis. Plants were transformed with a construct (GU-Ds-US) containing a Ds (Dissociation) transposable element inserted between two partially deleted GUS reporter gene segments. Homologous recombination between the overlapping GUS fragments generates clonal sectors visible upon staining for GUS activity. Plants containing the GU-Ds-US construct and a source of Ac (Activator) transposase showed an over 1000-fold increase in the incidence of recombination relative to plants containing the same construct but lacking transposase. Transposon-induced recombination was observed in vegetative and floral organs, and several germinally transmitted events were recovered. Transposon-induced recombination appears to be a general phenomenon in plants, and thus may have contributed to genome evolution by inducing deletions between repeated sequences.

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Section: Detection Of Somatic Recombination Events In Transgenic Plantssupporting

confidence: 58%

Section: Discussionmentioning

confidence: 99%

Intrachromosomal homologous recombination in Arabidopsis induced by a maize transposon

Xiao

Peterson

2000

Mol Gen Genet

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“…Many attempts to investigate DNA recombination in plants use overlapping mutant copies of a bacterial reporter gene, introduced either as transient (33)(34)(35)(36)(37)(38)(39)(40) or stable transgenes (41)(42)(43)(44)(45), to provide measures of somatic DNA recombination leading to a functional copy of the bacterial transgene. Using similar bacterial transgenes, two reports (46,47) measured the frequency of meiotic DNA recombination (6 ϫ 10 Ϫ6 and Ͻ2 ϫ 10 Ϫ5 , respectively).…”

Section: Discussionmentioning

confidence: 99%

Rare germinal unequal crossing-over leading to recombinant gene formation and gene duplication in Arabidopsis thaliana

Jelesko

Harper

Furuya

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

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Small, multigene families organized in a tandem array can facilitate the rapid evolution of the gene cluster by a process of meiotic unequal crossing-over. To study this process in a multicellular organism, we created a synthetic RBCSB gene cluster in Arabidopsis thaliana and used this to measure directly the frequency of meiotic, intergenic unequal crossing-over between sister chromatids. The synthetic RBCSB gene cluster was composed of a silent ⌬RBCS1B::LUC chimeric gene fusion, lacking all 5 transcription and translation signals, followed by RBCS2B and RBC3B genomic DNA. Expression of luciferase activity (luc ؉ ) required a homologous recombination event between the ⌬RBCS1B::LUC and the RBCS3B genes, yielding a novel recombinant RBCS3B͞ 1B::LUC chimeric gene whose expression was driven by RBCS3B 5 transcription and translation signals. Using sensitive, single-photon-imaging equipment, three luc ؉ seedlings were identified in more than 1 million F2 seedlings derived from self-fertilized F1 plants hemizygous for the synthetic RBCSB gene cluster. The F2 luc ؉ seedlings were isolated, and molecular and genetic analysis indicated that the luc ؉ trait was caused by the formation of a recombinant chimeric RBCS3B͞1B::LUC gene. A predicted duplication of the RBCS2B gene also was present. The recombination resolution break points mapped adjacent to a region of intron I at which a disjunction in sequence similarity between RBCS1B and RBCS3B occurs; this provided evidence supporting models of gene cluster evolution by exon-shuff ling processes. In contrast to most measures of meiotic unequal crossing-over that require the deletion of a gene in a gene cluster, these results directly measured the frequency of meiotic unequal crossingover (Ϸ3 ؋ 10 ؊6 ), leading to the expansion of the gene cluster and the formation of a novel recombinant gene.Genome organization can directly affect the evolution of a gene. Single-copy genes or dispersed members of a multigene family evolve independently. In contrast, members of a multigene family organized as a gene cluster can exploit this organization to generate further gene duplications and novel recombinant genes by a process of unequal crossing-over. For example, a single intergenic unequal crossover event in a gene cluster results in four simultaneous alterations: a deletion, a duplication, and two reciprocal, recombinant genes. The impact of such unequal crossover events evidently have been important in the evolution of complex loci such as HOX (1), amylase (2), globin (3), MHC (4), Ig (5), the maize R-r complex (6), RBCS (7), and plant disease-resistance loci (8-10). Although DNA sequencing of gene clusters provides information about past changes in a particular gene cluster, it can only estimate the rate of unequal crossing-over in terms of geological time scales.In multicellular organisms, unequal crossing-over is implicated in several genetic disorders. This was demonstrated first with the Drosophila bar locus (11). Subsequent research with the bobbed locus demonstrated ...

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“…al. 1996;Shalev et al 1999) or to induce DNA doublestrand breaks (DSBs) using chemicals or transposons in plants (Lowe et al 1992;Chiurazzi et al 1996;Shalev and Levy 1997;Xiao and Peterson 2000). Nevertheless, thus far little is known about HR in higher plants.…”

Section: Introductionmentioning

confidence: 99%

Enhanced homologous recombination caused by the non-transcribed spacer of the rDNA in Arabidopsis

et al. 2001

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The problem of the low frequency of homologous recombination observed in higher plants has been approached in several ways. Here, we report a new strategy to enhance homologous recombination in Arabidopsis. In Escherichia coli and Saccharomyces cerevisiae, hotspots that enhance homologous recombination nearby have been identified in regions close to sites associated with the blocking of DNA replication forks or with intensive transcriptional activity. In yeast, a recombination hotspot (HOT1) was found in a region spanning two non-transcribed spacers (NTS) between ribosomal RNA genes (rDNA), which contains both a replication fork barrier ( RFB) and the promoter for transcription of the 35S rRNA gene. Since rDNA has a common structure among eukaryotes, we analyzed the effect of the endogenous NTS on homologous mitotic recombination in a higher plant. We constructed transgenic lines of Arabidopsis containing this NTS and a recombination substrate, in which two 3'- and 5'-deleted uidA (beta-glucuronidase) genes with partially overlapping sequences are separated by a Hyg(r) gene. Reconstitution of functional uidA genes by homologous recombination was monitored by histochemical GUS staining. We found that recombination occurred more frequently in all organs tested in F (Fork block) lines transgenic for the NTS than in C (Control) lines without the NTS. The average number of GUS+ spots on leaves in F lines was more than nine-fold higher than in C lines. Furthermore, by genomic Southern analysis, post-recombinational molecules were detected in a transgenic line, F43, which had an extremely high number of GUS+ blue spots. These results strongly suggest that NTS-dependent enhancement of homologous recombination may be a common feature of higher plants as well as yeast.

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Enhancement of Somatic Intrachromosomal Homologous Recombination in Arabidopsis by the HO Endonuclease

Cited by 17 publications

References 35 publications

Intrachromosomal homologous recombination in Arabidopsis induced by a maize transposon

Intrachromosomal homologous recombination in Arabidopsis induced by a maize transposon

Rare germinal unequal crossing-over leading to recombinant gene formation and gene duplication in Arabidopsis thaliana

Enhanced homologous recombination caused by the non-transcribed spacer of the rDNA in Arabidopsis

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