Deficiency analysis of megagametogenesis
--Erik Vollbrecht

By crossing together appropriately chosen reciprocal translocation stocks, one can construct euploid genotypes that, after meiosis, segregate (haploid) spores containing relatively small interstitial deficiencies (for discussion, see Birchler's chapter on segmental aneuploidy in The Maize Handbook.). Since haploid spores are the predecessors of the gametophytes (pollen and embryo sac [ES]), and deficient genotypes rarely transmit through either pollen or ES, gametophyte-specific functions are probably encoded by loci within such deficiency-defined regions. By determining the gametophytic defects conferred by deficiency for a particular region, one can infer the function(s) of gametophyte-expressed genes within that region. Thus, I am using translocation heterozygotes that segregate (balanced:duplicate:deficient) spores in a ratio of (2:1:1) to determine the affects of various loss of function genotypes on ES development. Stocks for segmental analysis of 1L and 3L were kindly supplied by Jim Birchler. The Stock Center supplied the Tp9; Df3 stock, which was described in some detail by M. M. Rhoades, MM (in Replication and Recombination of Genetic Material, Peacock and Brock, eds., 1968). The Tp9; Df3 stock also segregates 25% deficient spores, which do not transmit through either gametophyte due to deficiency for ~10% of 3L. Immature ears were fixed and processed for analysis by confocal laser scanning microscopy (CLSM). Deficiency phenotypes are inferred from the frequency of occurrence of novel ES phenotypes in a population from a single ear. Analysis is still in progress, but phenotypes observed thus far include ESs that develop to the 2-nucleate stage and persist as immature ESs, ESs that degenerate precociously, 2-nucleate ESs with non polar nuclear distribution, and 4-nucleate ESs with nuclear degeneration at the micropylar pole but not the chalazal pole. Initial observations suggest that ESs containing the corresponding duplicate genotypes, which also segregate at 25% on the ears analyzed but transmit with regularity, may develop slower than do their euploid "siblings". 

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