A proposed method to determine the time of occurrence of nondisjunction induced by the r-X1 deficiency

The r-X1 deficiency is an X-ray induced submicroscopic deletion of the R locus on chromosome 10. When deficiency-bearing gametes are fertilized, a large percentage of the resultant embryos are aneuploid. Because there is non-correspondence between embryo and endosperm markers, and because approximately equal numbers of trisomic and monosomic embryos are produced, it appears that the r-X1 deficiency induces chromosomal nondisjunction after meiosis is completed during the embryo sac (megagametophyte) divisions (Weber, in Maize for Biological Research, p. 79, 1982). It is not known if the nondisjunctive event is restricted to one specific post-meiotic division.

The following assumptions are used in this discussion: a) only one nondisjunctional event occurs during the development of the embryo sac, b) the two nuclei move to opposite poles after the first megagametophyte division, c) after the third division any one of the four nuclei at each pole has an equal probability to migrate to the center of the embryo sac to become a polar nucleus, and d) any one of the four nuclei at the micropylar end has an equal probability to form the egg.

If nondisjunction occurs during the first megagametophyte division, all eggs will be aneuploid, one polar nucleus will be nullisomic (n-1), and the other polar nucleus will be disomic (n+1). When the two polar nuclei are fertilized by a haploid pollen grain, euploid endosperm will invariably be produced. If nondisjunction occurs at the second megagametophyte division, all nuclei at one pole of the embryo sac will be euploid and all nuclei at the other pole will be aneuploid. When nuclei migrate from each pole of the mature embryo sac to form the polar nuclei, one polar nucleus will invariably be aneuploid and the other euploid; thus, endosperm formed from such an embryo sac will invariably be aneuploid. Half of the embryos would be aneuploid. Finally, if nondisjunction occurs at the third division, all the nuclei at one pole will be euploid and half of the nuclei at the other pole will be aneuploid, and half of the kernels formed from such embryo sacs will have aneuploid endosperm. One fourth of the embryos would be aneuploid.

If one selects kernels that are monosomic for a specific chromosome, one can predict the chromosomal constitution of the endosperm assuming that nondisjunction occurred at specific divisions. If nondisjunction occurred at the first division, the endosperm invariably would be euploid. If nondisjunction occurred at the second division, the endosperm invariably would be aneuploid for the monosomic chromosome. Half of the time the endosperm would be hypoploid and half of the time it would be hyperploid. If nondisjunction occurred at the third division, the endosperm would be aneuploid for the monosomic chromosome half of the time and euploid half of the time.

Thus, the division at which nondisjunction occurs can readily be determined if a suitable genetic marker can be utilized which expresses clearly-defined dosage effects in the endosperm. Endosperm of kernels monosomic for this chromosome could be analyzed. A suitable gene for this purpose would be the Y locus on chromosome 6 which controls the level of carotenes in the endosperm of the kernel. Kernels homozygous for a particular recessive allele of this locus, y-pastel-8549, have white endosperm and produce pale-green seedlings when germinated at 37 C (Robertson, MNL 34:73). We have crossed r-X1 bearing plants which were R/r-X1; Y/Y by male parents that were r/r; Y/y-pas. The purple (R/r) kernels will be discarded and the anthocyaninless (r/r-X1) kernels will include kernels with embryos that are monosomic for chromosome 6. Monosomic 6 embryos that are y-pas/- will give rise to pale-green seedlings when germinated at 37 C. Root-tip cells from such seedlings will contain 19 chromosomes and only one satellited chromosome (chromosome 6). Endosperm samples from such kernels obtained prior to germination will be analyzed for their levels of carotenes by high performance liquid chromatography (HPLC). We have analyzed the levels of carotenes in kernels that are Y Y Y, Y Y y, Y y y, or y y y with HPLC, and we have found that it is possible to distinguish unambiguously the different kernel types. As expected, we find a strict dosage relationship between the number of dominant alleles and the amount of carotene present. Single kernel analysis by this method appears to be quite feasible. The data obtained will allow the precise determination of the number of Y alleles present (which is the number of chromosome 6's contributed by the maternal parent) in kernels monosomic for chromosome 6.

If nondisjunction of chromosome 6 takes place at the first megagametophyte division, the endosperm of kernels with monosomic 6 embryos will invariably contain two Y alleles. If it takes place at the second division, these kernels will have one or three Y alleles in their endosperm. If it takes place at the third division, half of the kernels will have two, a fourth will have one, and a fourth will have three Y alleles in their endosperm. In this way we hope to precisely define the division at which the r-X1 induced nondisjunctional event takes place.

J. D. Shadley, K. Simcox and D. F. Weber

Please Note: Notes submitted to the Maize Genetics Cooperation Newsletter may be cited only with consent of the authors.

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