Rare germinal unequal crossing-over leading to recombinant gene formation and gene duplication in Arabidopsis thaliana.
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 DeltaRBCS1B::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 DeltaRBCS1B::LUC and the RBCS3B genes, yielding a novel recombinant RBCS3B/ 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 RBCS3B/1B::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-shuffling 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 crossing-over (approximately 3 x 10(-6)), leading to the expansion of the gene cluster and the formation of a novel recombinant gene.
Jelesko
J G
JG
Department of Plant and Microbial Biology, 111 Koshland Hall, University of California, Berkeley, CA 94720-3102, USA.
Harper
R
R
Furuya
M
M
Gruissem
W
W
D017360:Arabidopsis / Q000235:genetics*; D001483:Base Sequence; D003434:Crossing Over, Genetic; D017931:DNA Primers; D005091:Exons; D020440:Gene Duplication; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D005813:Genes, Synthetic; D008969:Molecular Sequence Data; D005810:Multigene Family; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000235:genetics; D011995:Recombination, Genetic; D012273:Ribulose-Bisphosphate Carboxylase / Q000235:genetics*; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D012273:Ribulose-Bisphosphate Carboxylase