II. 7. Microspore derived chromosome number and structural variants of barley (Hordeum vulgare L.).
Wolfgang Friedt and Bärbel Foroughi-Wehr, Department of Plant Genetics, Gesellschaft für Strahlen- und Umweltforschung mbH München, D-8059 Grünbach, FRG.
In previous investigations it was demonstrated that a great variation of chromosome number may occur in somatic cells of microspore derived plants in various species (D'Amato 1977). In root tips of regenerated barley plants haploid, diploid, tetraploid as well as aneuploid cells were observed (Mix et al. 1978). The present study deals with the nature of chromosomal variation in the germ cells (PMCs) of microspore derived barley plants.
Diploid and tetraploid barley stocks were used as donors for anther culture. As a diploid donor the cultivar 'Dissa' was grown. This variety proved to respond best to the culture methods applied. As tetraploid donors the cv. 'Haisa II' and a selected strain out of crosses of 'Haisa II' to other tetraploid stocks were used. This material was originally chosen in order to detect obscure chromosome mutations in the tetraploids. The culture methods and staining techniques applied were described earlier (Foroughi-Wehr et al. 1976; Friedt 1979).
A. Anther culture of diploid donor plants
Anther derived progenies show large variation in fertility. Not only in the first (A1) but also in successive generations, completely sterile plants occur.
A detailed cytogenetic study of a random sample of 280 out of 2000 plants obtained through anther culture of barley cv. 'Dissa' revealed that the chromosome numbers in germ cells do not vary as much as in somatic cells. Only haploid, diploid and tetraploid but no aneuploid complements have been found in PMCs so far and their frequencies have been estimated as approximately 20%, 70% and 10%, respectively (Table l).
Table l. Progeny of anther culture of barley cv. 'Dissa' in the Al-generation*
The diploid progenies usually show complete seed set, whereas the tetraploid individuals exhibit a large variation in fertility ranging from 0 to almost 100%. The haploid progenies are usually sterile. However, some spikes were found with one single seed. This can be explained by premeiotic fusion or endoreduplication of haploid cells and/or by meiotic C-metaphase formation leading to di-haploid gametes (Fig. 1A).
Figure 1. PMCs from haploid (A), diploid (B, C,
D) and tetraploid (E, F) individuals.
A. PMC with 14 univalents produced by premeiotic fusion or endoreduplication as an explanation for occurrence of single seeds in spikes of haploid individuals.
B. PMC in MI with seven bivalent-rings (7 II).
C. Heterozygous reciprocal translocation obtained through anther culture of tetraploid barley. PMC in MI with five bivalent-rings (5 II) and a ring of four chromosomes (1 IV).
D. Two PMCs in AI with normal chromosome-separation (left) and chromatid bridge-formation (right), respectively.
E. PMC in MI with 6 II and 4 IV.
F. PMC in AI with irregular separation of chromosomes (15:13) explaining the production of a trisomic plant via anther culture of tetraploid barley.
In other cases plants with more than 20 fully sterile ears spontaneously produced one completely fertile spike. In addition, one plant with diploid and tetraploid ears was also found. This may be explained with the presence of diploid and tetraploid cells in the somatic tissue of the Plants. A number of regenerated individuals showed extreme variation in phenotypic characters such as chlorophyll defects, stunted growth and leaf irregularities.
All of the diploid plants studied except one were cytologically normal. The latter proved to be heterozygous for a reciprocal translocation.
B. Anther culture of tetraploid donor plants
Out of 23000 cultured anthers isolated from tetraploid stocks of barley 32 white and 19 green plants were regenerated. Most of the latter were phenotypically similar to normal diploid barley. However, in meiotic first anaphase of two diploid individuals chromatid-bridge formation was observed (Fig. 1D). One of the two plants showed in addition varying chromosome numbers in the PMCs. Another plant was trisomic (2n=2x+1=15) for chromosome 6 or 7 (nucleolar chromosomes; Friedt et al. 1977), three were heterozygous for reciprocal translocations (Fig. 1C), four individuals were tetraploid (2n=4x=28; Fig. E, F), and the other plants showed normal meiotic chromosome behavior (Fig. 1B). The progenies of successive generations will be investigated for variation in quantitative characters. Additionally, backcrosses to the original diploid barley cv. 'Haisa II' (already carried out) will reveal further possible hidden homozygous chromosome mutations.
The present study shows that only about 20% of anther derived barley plants are haploid. However, together with the approximately 70% diploids which are produced via spontaneous diploidization 90% of the anther progeny can be used immediately in barley breeding programs. The remaining 10% which are obviously exclusively tetraploids can be useful in breeding programs for tetraploid barley as well as for genetic studies.
Because of the occurrence of chromosome mutations in diploid progenies derived from anther culture of tetraploid plants it can be assumed that also in the spontaneous diploid progenies from anther culture of diploid donor plants hidden chromosome-as well as gene-mutations may be present. Such obscure mutations would be disadvantageous in breeding programs. To find out the nature and rate of such mutations, the anther progeny have been backcrossed to their original diploid donor variety. The first backcross generation is going to be investigated cytogenetically.
We wish to thank Dr. Gisela Gosch-Wackerle for very helpful advice.
D'Amato, F. 1977. Cytogenetics of differentiation in tissue and cell cultures. In J. Reinert and Y.P.S. Bajaj (Ed.), Applied and fundamental aspects of plant cells, tissue and organ culture, 343-356. Springer-Verlag, Berlin.
Foroughi-Wehr, B., G. Mix, H. Gaul and H. M. Wilson. 1976. Plant production from cultured anthers of Hordeum vulgare L. Z. Pflanzenzüchtg. 77:198-204.
Friedt, W. 1979. Untersuchungen an autotetraploiden Gersten unter besonderer Berücksichtigung der Diploidisierung II. Z. Pflanzenzüchtg. 82:311-339.
Friedt, W., B. Foroughi-Wehr, G. Mix and H. M. Wilson. 1977. Anther culture of autotetraploid Hordeum vulgare and the production of trisomic individuals. BGN 7:29-33.
Mix, G., H. M. Wilson and B. Foroughi-Wehr. 1978. The cytological status of plants of Hordeum vulgare L. regenerated from microspore callus. Z. Pflanzenzüchtg. 80:89-99.
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