Plant materials: Can 7DL was derived from the misdivision of chromosome 7D in Can monosomic 7D that had been produced by a cross between Can8 and Stewart durum wheat. We performed eight-time successive backcrosses of Can ditelosomic 7DL to CS. Finally through crossing BC8 plants with nulli7D-tetra7B and selfing the resulting plants, 20 offspring with 40 normal chromosomes and a pair of 7DL were isolated. For cytologically identifying 7DL, BC plants were crossed to CS ditelo 7DS and the chromosome constitution (41+2t, 20"+1t1''') was confirmed in every BC generation.
Cytological analysis: Root tips were pretreated in ice water for 24 hours and fixed with fresh Farmer's fluid. Anthers at metaphase-I were fixed in Farmer's fluid.Chromosome preparations were Made by the acceto-carmine squash method.
alpha-amylase isozymes: The genes for several alpha-amylase isozymes are located on the respective long arms of six chromosomes belonging to homoeologous groups 6 and 7 of CS (Nishikawa and Nobuhara 1971). We used this isozyme as a genetic marker for the identification of ditelosomic 7DL. The same zymogram technique described by Nishikawa et al (1979) was used for alpha-amylase of germinating seeds in this study.
Test-crossing with ditelosomic 7DS: Plants with alpha-amylase isozyme pattern specific to ditelosomic 7DL were crossed to ditelosomic 7DS. If the plants are authentic ditelosomic 7DL, the chromosome pairing in the F1 is 20''+2t'. If they are ditelosomic 7DS, the F1 shows 20"+t''. If they are neither ditelosomic 7DL nor 7DS, the F1 shows 19"+2(1t'').
Results and discussion
In all 20 developed lines, somatic chromosome constitution was 2n=40+2t
in root tip cells and chromosome pairing was 20''+t'' in pollen mother cells
(Table 1). Considering the production process, the
developed lines were either ditelosomic 7DL or 7DS. The difference in length
between the long and short arms of chromosome 7D is small (Furuta et al. 1988,
Endo and Gill 1996).
One of the genes for alpha-amylase isozymes is located on 7DL (Nishikawa and Nobuhara 1971). The zymogram of ditelosomic 7DL should be identical to that of euploid CS but that of ditelosomic 7DS lacks the band of 7DL (Fig. 1). Sixteen out of the 20 lines were ditelosomic for 7DL and four lines were ditelosomic for 7DS (Table 1). Of these 16 lines, three lines were randomly selected and crossed to ditelosomic 7DS. The F1's all showed meiotic chromosome configuration 20"+2t'(Table 2, Fig. 2).
Height and seed fertility of these CS ditelosomic 7DL plants were intermediate between the euploid CS and ditelosomic 7DS (Table 3). The ditelosomic 7DL plants were cytologically stable, and their spike morphology was normal.
CS is the most important variety among wheat lines to chromosome engineering. Since Sears (1954) established the monosomic series of CS, an array of aneuploid lines has been produced in this variety. Sears and Sears (1978) tried to produce ditelosomic 7DL using 2000 seeds but failed to recover the ditelosomic 7DL. Ditelosomic 7DL stock of CS is currently being extracted at John Innes Centre from the double ditelosomic 7D stock (Devos et al. 1999). The principal use of ditelosomic lines is to provide cytologically recognizable chromosomes in identifying aneuploids, locating and mapping genes, and determining degrees of meiotic pairing and somatic association. The CS ditelosomic 7DL stock developed by us in the present study will be useful for analyzing genes on chromosome 7D.