The isolation and characterization of Fcu revertants
No full colored germinal revertants have been reported from the Fcu controlling element system (Gonella and Peterson, Molec Gen Genet 167:29–36, 1978). Of 30 full colored kernels isolated from crosses involving the Fcu responsive haplotype r1-cu and Fcu, all turned out to be nonrevertant, and had appeared to be full colored due to the variable high background of aleurone color expression by r1-cu. We recently found that additional r1 haplotypes respond to Fcu, and some of these haplotypes have a paler aleurone color background than r1-cu (Stinard, MNL 77:77–79, 2003). We proposed that mutability, and therefore reversion events, in the Fcu system is most likely due to changes at Fcu and not changes at the r1 locus. In order to isolate Fcu revertants for further study, we made crosses of an r1-g Fcu line onto a responsive pale r1 haplotype, R1-r(Venezuela559-PI302355). Many full colored putative single kernel revertants were isolated from such crosses, and are summarized in Table 1. Many stable pale kernels that might represent Fcu losses were also observed, but these were not counted separately. Kernel counts are grouped by Fcu male parent because the possibility exists that multiple kernel revertants in such crosses could arise from revertant tassel sectors and might represent a single reversion event. If one counts each putative revertant as an independent event, the frequency of reversion observed in this population is 3.0 × 10−3. However, if one considers multiple revertant kernels arising from one male parent as single events, the reversion rate becomes 8.1 × 10−4. These two frequencies provide logical bounds for the rate of Fcu reversion in this population. All of these putative revertants need to be grown out and tested for the presence of contamination markers (y1 and wx1 from the Fcu parent) in order to rule out the possibility of contamination. However, if any of these putative revertants are real, it is clear that reversion is occurring at Fcu (or some other locus that interacts with Fcu) and not at the r1 locus since the r1-g haplotype in the Fcu parent is not mutable in the presence of Fcu.
One of the Fcu male parents (2003-2653-6) used in these crosses was found to be heterozygous for a reversion event that must have arisen unnoticed in the previous generation. Kernel counts from this male parent are summarized in Table 2. Outcrosses of this particular parent to the R1-r(Venezuela 559-PI302355) tester resulted in 1:1 segregation for full colored and sectored/pale kernels. The self-pollinated ear from the male parent plant was homozygous for the y1 and wx1 contamination markers, ruling out contamination of the Fcu parent line. The reproducibility of the 1:1 segregation in four separate outcrosses rules out contamination arising during pollination, as well as contamination occurring in the R1-r(Venezuela 559-PI302355) parent line. This event is the first proven instance of reversion in the Fcu system, and the revertant has been named Fcu-R2003-2653-6. Crosses are planned to determine whether Fcu-R2003-2653-6 maps to the same chromosomal location as Fcu, or whether it represents a transpositional event or a change at an unlinked locus. The other putative single kernel revertants will be subjected to the same analysis if they prove not to be contaminants.
Table 1. Counts of full colored putative single kernel revertants from the cross R1-r(Venezuela559-PI302355 fcu × r1-g Fcu wx1 y1.
|Fcu male parent||female parent||No. sectored and pale kernels||No. full colored kernels|
Frequency of reversion = 3.0 × 10−3
Table 2. Counts of kernels from crosses involving the heterozygous Fcu revertant parent 2003-2653-6. Test cross: R1-r(Venezuela559-PI302355 fcu × r1-g; Fcu/Fcu-R; wx1 y1.
|female parent||No. sectored and pale kernels||No. full colored kernels|
1:1 χ2 = 0.935 (not significant)