title: High-resolution tyramide-FISH mapping of markers tightly linked to the male-fertility restoration (Ms) locus of onion.
abstract: Tyramide FISH was used to locate relatively small genomic amplicons from molecular markers linked to Ms locus onto onion chromosome 2 near the centromere, a region of relatively low recombination. Fluorescence in situ hybridization (FISH) has not been readily exploited for physical mapping of molecular markers in plants due to the technical challenge of visualizing small single-copy probes. Signal amplification using tyramide (tyr) FISH can increase sensitivity up to 100-fold. We used tyr-FISH to physically locate molecular markers tightly linked to the nuclear male-fertility (Ms) restoration locus of onion onto mitotic metaphase, pachytene, and super-stretched pachytene chromosomes. Relatively short genomic amplicons (846-2251 bp) and a cDNA clone (666 bp) were visualized in 9-42 % of observed cells. The markers were assigned to proximal locations close to the centromere on the long arm of chromosome 2, a region of lower recombination, revealing that tightly linked markers may be physically distant from Ms. This result explains why several labs have identified molecular markers tightly linked to the Ms locus after screening relatively few DNA clones or primers and segregating progenies. Although these markers are still useful for marker-aided selection, our results indicate that map-based cloning of Ms will likely be difficult due to reduced recombination near this gene.
authors.affiliation: Center of Molecular Biotechnology, Department of Genetics and Biotechnology, Russian State Agrarian University-Timiryazev Agricultural Academy, 49, Timiryazevskaya Str., 127550, Moscow, Russia. ludmila.khrustaleva19@gmail.com.
authors.forename: Ludmila
authors.initials: L
authors.lastname: Khrustaleva
authors.affiliation: Department of Horticulture, University of Wisconsin, Madison, WI, 53706, USA.
authors.forename: Jiming
authors.initials: J
authors.lastname: Jiang
authors.affiliation: USDA-ARS and Department of Horticulture, University of Wisconsin, Madison, WI, 53706, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Havey
mesh_terms: D002503:Centromere / Q000235:genetics; D032461:Chromosomes, Plant / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D005819:Genetic Markers; D017404:In Situ Hybridization, Fluorescence; D019697:Onions / Q000235:genetics*; D020161:Physical Chromosome Mapping / Q000379:methods*; D051479:Plant Infertility / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D020641:Polymorphism, Single Nucleotide
chemical_list: D018076:DNA, Complementary; D018744:DNA, Plant; D005819:Genetic Markers