BGN 2: Barley haploid studies BARLEY GENETICS NEWSLETTER, VOL. 2, II. RESEARCH NOTES
Kasha et al., pp. 36-41

II.10. Barley haploid studies.

K. J. Kasha, E. Reinbergs, W. A. Johns, N. C. Subrahmanyam and K. M. Ho. Department of Crop Science, University of Guelph, Guelph, Ontario, Canada.

The barley genetic studies at Guelph are closely associated with the breeding program and the haploid project is one good example of this. The cytogenetic studies (K. J. Kasha) are being conducted on materials from the breeding program (E. Reinbergs). Thus, the doubled haploids resulting from genetic and cytological studies of haploid formation and development are being evaluated for potential varieties or for obtaining information on breeding techniques. Following are three brief outlines of thesis projects that are in progress. In addition, Dr. E. J. Walsh has just begun a Post-doctorate Fellowship project in the area of breeding techniques with the doubled haploid materials.

1. Haploids in barley breeding.  W. A. Johns
The purpose of this study is to compare haploids with the bulk and pedigree methods of barley breeding for both qualitative and quantitative characters. Homozygous doubled haploid lines are being produced from F1 hybrids (Dr. Kasha's program) and will be compared under field conditions with the parental cultivars and recommended commercial varieties. Materials from the same crosses will be handled by the bulk and pedigree methods.

Preliminary field observations on doubled haploids in 1970 and 1971 revealed the striking uniformity of the lines. Qualitative characters were simple to detect. In 1971, a total of 65 doubled haploid lines from two crosses were grown in single-row 10' plots along with randomly selected F3's. Parental cultivars were placed as checks in every 3rd row.

Frequencies of lines with rough vs. smooth awns, two-rowed vs. six-rowed spikes and disease resistance indicated that a random sample of genotypes was obtained following "gamete selection" from F1 hybrids. Sufficient seed from 19 unselected doubled haploids permitted a replicated yield test to be run in 1971. Four of the doubled haploids in this test significantly outyielded the parental lines and two other commercially recommended varieties that were included. Other quantitative characters have not been analyzed as yet.

In 1972 we expect to have a total of 400 doubled haploids from three different crosses under field evaluation. In addition, the F4 rows and bulks from F3 rows will be grown. Replicated yield tests will be conducted to compare the most promising doubled haploids with parental lines and commercial varieties.

2. A cytological study of chromosome elimination during haploid formation in barley    N. C. Subrahmanyam

Numerous haploids of cultivated barley have been produced from the cross of Hordeum vulgare (2n=14) by H. bulbosum (2n=14), Dihaploids (2n=14) can also be produced by crossing the tetraploid (2n=28) cytotypes of vulgare and bulbosum. However, the cross diploid vulgare (2n=14) by tetraploid bulbosum (2n=28) usually gives a stable triploid hybrid (2n=21).

Reports of Kao and Kasha (1969), Kasha and Kao (1970), Symko (1969) and Lange (1969, 1971) independently suggested the hypothesis of chromosome elimination as a mechanism leading to haploidy in barley. Kasha and Kao (1970) suggested that the genome balance between the parental species was important in chromosome stability. The present study aims to trace the process of chromosome elimination in tissues (embryo and endosperm) with different genomic constitutions, The following table presents different cross combinations and assuming fertilization, the expected genomic constitutions of embryo and endosperm which are being used for cytological examinations:

Table 1.

Squash preparations of embryo and endosperm are being made to determine the frequencies of cells with different chromosome numbers and abnormalities. Table 2 presents mainly chromosome variations in the embryos of VB constitution. The process of chromosome elimination appears to be gradual and the elimination can occur at any time after fertilization. The percentage of cells with 7 chromosomes (haploid set) increases with the age of the embryo while the percentage of abnormalities (extranuclear material, dense staining bodies, etc,) decrease. From the same cross endosperm cells with the expected VVB genomic constitution were quite variable in chromosome number, ranging from 14 to 28.

Table 2. Chromosome variations and abnormalities in embryos with VB constitution.

In embryos from a reciprocal cross between the diploid parents a similar pattern of chromosome elimination was observed. However, in this cross, the endosperm with BBV constitution appeared to have cells with relatively few abnormalities and with more stable chromosome numbers. Even at 7 days, most dividing cells had 21 chromosomes.

Embryos VBB genomic constitution were mostly stable and all the seedlings obtained from this cross combination had stable chromosome numbers (2n=21).

Thus it is quite evident that chromosome or genome balance in the hybrids is important. Furthermore, techniques which would speed up the elimination of bulbosum chromosomes might lead to higher frequencies of haploids following embryo culture.

References:

Kao, Ko N., and Kasha, K. J. 1969. Haploidy from interspecific crosses with tetraploid barley. Proc. 2nd Int. Barley Genet. Symp. II: 82-88.

Kasha, K. J. and Kao, K. N. 1970. High frequency haploid production in barley (Hordeum vulgare L.). Nature 225:874-876.

Lange, W. 1969. Cytogenetical and embryological research on crosses between Hordeum vulgare and H. bulbosum. Versl. landbowk. Onderz. 719:162.

Lange, W. 1971. Crosses between Hordeum vulgare L. and H. bulbosum L. II. Elimination of chromosomes in hybrid tissues. Euphytica 20:181-194.

Symko, S. 1969. Haploid barley from crosses of Hordeum bulbosum (2x) X H. vulgare (2x). Can. J. Genet. Cytol. 11:602-608.

3. Genetic control of chromosome elimination during the formation of barley haploids.   K. M. Ho

The results from all the possible interspecific crosses between H. vulgare and H. bulbosum (both diploid and tetraploid forms) revealed that to obtain chromosome stable hybrids the genomes of H. vulgare and H. bulbosum should be in a 1 to 2 ratio (Kao and Kasha, 1970). However, a hybrid with 27 chromosomes was occasionally produced from the cross between tetraploid forms of these two species, indicative that a specific chromosome(s) may carry factor(s) controlling chromosome stability in these interspecific hybrids (Kasha et al., 1970).

To test this hypothesis, crosses between the seven primary trisomic types of diploid H. vulgare and the autotetraploid form of H. bulbosum are being carried out.

Such crosses are expected to produce two types of hybrid progeny, a) 21 chromosome triploid hybrids and b) 22 chromosome hybrids (3x+1) when the trisomic chromosome is transmitted. The frequency of 22 chromosome hybrids should be similar to the trisomic transmission frequency in diploid vulgare. If specific chromosomes carry a factor(s) controlling chromosome stability, the 22 chromosome hybrids will not occur or be rare in crosses with plants trisomic for that chromosome. In such instances one might expect to find 8 chromosome haploids (X+1) if they are able to survive.

The preliminary results of crosses between primary trisomics of H. vulgare and the tetraploid form of H. bulbosum are given in Table 3. The results show that the frequency of 22 chromosome hybrid progeny is low from crosses involving chromosomes 2 and 3, being 1.8 and 5.3% respectively. However, 8 chromosome progeny have not been found to date.

Table 3. Progeny from crosses between primary trisomics of diploid H. vulgare and tetraploid H. bulbosum.

It would appear that chromosome 2 and possibly chromosome 3 contain major genetic factors which are critical to the chromosome balance and stability in the interspecific hybrids between H. vulgare and H. bulbosum. Additional crosses using telotrisomics for chromosomes 2 and 3 will be conducted.

References:

Kao, K. N., and K. J. Kasha. 1970. Haploidy from interspecific crosses with tetraploid barley. Barley Genetics II:82-89. Kasha, K. J., K. N.

Kao and E. Reinbergs. 1970. Genetic control over chromosome stability in hybrids from interspecific Hordeum crosses. Genetics 64(2):33s.

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