Inheritance and mapping of 2n-egg production in diploid alfalfa

Abstract

The production of eggs with the sporophytic chromosome number (2n eggs) in diploid alfalfa (Medicago spp.) is mainly associated with the absence of cytokinesis after restitutional meiosis. The formation of 2n eggs through diplosporic apomeiosis has also been documented in a diploid mutant of M. sativa subsp. falcata (L.) Arcang. (2n = 2x = 16), named PG-F9. Molecular tagging of 2n-egg formation appears to be an essential step towards marker-assisted breeding and map-based cloning strategies aimed at investigating and manipulating reproductive mutants of the M. sativa complex. We made controlled crosses between PG-F9 and three wild type plants of M. sativa subsp. coerulea (Less.) Schm. (2n = 2x = 16) and then hand-pollinated the F1 progenies with tetraploid plants of M. sativa subsp. sativa L. (2n = 4x = 32). As a triploid embryo block prevents the formation of 3x progenies in alfalfa because of endosperm imbalance, and owing to the negligible selfing rate, seed set in 2x-4x crosses was used to discriminate the genetic capacity for 2n-egg production. F1 plants that exhibited null or very low seed sets were classified as normal egg producers and plants with high seed sets as 2n-egg producers. A bulked segregant analysis (BSA) with RAPD (random amplified polymorphic DNA), ISSR (inter-simple sequence repeat), and AFLP (amplified fragment length polymorphism) markers was employed to identify a genetic linkage group related to the 2n-egg trait using one of the three F1 progenies. This approach enabled us to detect a paternal ISSR marker of 610 bp, generated by primer (CA)8-GC, located 9.8 cM from a putative gene (termed Tne1, two-n-eggs) that in its recessive form determines 2n eggs and a 30% recombination genomic window surrounding the target locus. Eight additional RAPD and AFLP markers, seven of maternal, and one of paternal origin, significantly co-segregated with the trait under investigation. The minimum number of quantitative trait loci (QTLs) controlling seed set in 2x-4x crosses was estimated by ANOVA and regression analysis. Four maternal and three paternal independent molecular markers significantly affected the trait. A paternal RAPD marker allele, mapped in the same linkage group of Tne1, explained 43% of the variation for seed set in 2x-4x crosses indicating the presence of a major QTL. A map of the PG-F9 chromosome regions carrying the minor genes that determine the expression level of 2n eggs was constructed using selected RAPD and AFLP markers. Two of these genes were linked to previously mapped RFLP loci belonging to groups 1 and 8. Molecular and genetic evidence support the involvement of at least five genes.

A genetic linkage map for watermelon derived from a testcross population: (Citrullus lanatus var. citroides × C. lanatus var. lanatus) × Citrullus colocynthis

“…ISSR markers have been mainly used in genetic diversity studies (Levi and Rowland 1997;Prevost and Wilkenson 1999;Joshi et al 2000). However, a few recent studies employed ISSR markers in genetic mapping of crop plants ( Barcaccia et al 2000;Santra et al 2000;Casasoli et al 2001). Like the RAPD markers, the ISSR markers in the present study are highly reproducible from experiment to experiment following optimization of their annealing temperature (Table 2).…”

Genetic diversity and reproductive biology in ecotypes of the facultative apomict Hypericum perforatum L.

“…Inter-microsatellite markers were assayed using seven different ISSR primers (synthesized by Life Technologies, Inc.) anchored at the 3 0 or 5 0 terminus of the simple CA repeat and extended into the flanking sequence by two or three nucleotide residues (Table 2). The protocol used for detecting ISSR polymorphisms was that reported by Barcaccia et al (2000a). Photographs (DC120 camera, Kodak) of the polymerized DNA fragments were taken after staining of the 2% agarose gels with ethidium bromide.…”

A genetic linkage map of grape, utilizing Vitis rupestris and Vitis arizonica

“…Although ISSR markers have not been applied to grape mapping projects, they have been used successfully in mapping projects of other plant species (Barcaccia et al 2000;Winter et al 2000;Casasoli et al 2001). The ISSR procedure was faster and easier than the AFLP procedure, but less efficient with an average of four markers per ISSR primer.…”