Euphytica 29 (1980) 635-640

INTERFERING WITH REGULAR MEIOTIC BEHAVIOUR IN AVENA SATIVA AS A METHOD OF INCORPORATING THE GENE FOR MILDEW

RESISTANCE FROM A. BARBATA

HUGH THOMAS, W. POWELL and T. AUNG

Welsh Plant Breeding Station, Plas Gogerddan, nr Aberystwyth, Dyfed, SY23 3EB, Wales, UK

Received 25 February 1980

INDEX WORDS

Avena sativa, oat, Avena barbata, wild oat, mildew resistance, gene transfer, meiotic behaviour.

SUMMARY

The regular meiotic behaviour of the cultivated oat Avena sativu (2n = 6x = 42) is genetically controlled. The factors which control the diploid-like meiotic behaviour also restrict the amount of pairing that occurs between alien chromosomes and their homoeologues in A.sativa, and hence increases the difficulties of introducing desirable variation from wild species into the cultivated oat. A genotype of the diploid species A.longiglumis which interferes with the reguiar meiotic behaviour of A. sativa can be used to induce pairing between alien chromosomes and their corresponding chromosomes in A. sativa. Using this procedure the dominant geneconferringmildew resistance has been transferred from the tetraploid weed species A. barbata into the cultivated oat.

INTRODUCTION

The regular formation of 21 bivalents at metaphase I in A. sativa, together with disomic inheritance, clearly shows that chiasmate associations are confined to pairs of homo- logous chromosomes. There is strong evidence (GAUTHIER & MCGINNIS, 1968; RAJ- HATHY & THOMAS, 1972, 1974) that regular meiotic behaviour in A. sativa is genetically controlled by a system analogous to the Ph gene in Triticum aestivum (RILEY & CHAPMAN, 1958). The activity of these pairing genes also restricts synapsis between alien chromosomes and their corresponding chromosomes in A. sativa and therefore limits the scope for introducing alien variation into the germplasm of the cultivated oat. THOMAS et al. (1975) failed to obtain any recombinants in a large population of selfed progeny of a monosomic addition line, in which the chromosome of the tetra- ploid species A. barbata carrying the gene conferring mildew resistance, was added to the A. sativa complement. However, the gene for mildew resistance has been success- fully transferred to A. sativa by means of an irradiation induced translocation (AUNG, et al., 1977). A limitation of the use of induced translocations to effect gene transfers is the undesirable consequences of duplications and deletions that arise in such chromo- some exchanges.

The transfer of genes from related species into the cultivated wheat has been success- fully accomplished through inducing alien chromosomes to pair with their ho- moeologues in wheat by suppressing (RILEY et al., 1968) or deleting the function of the

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Ph gene (SEARS, 1977). The genetically induced gene transfers involve an exchange between homoeologous chromosomes which have similar gene sequences, and do not result in deletion/duplications as in irradiation induced translocations.

Since the chromosome(s) on which the pairing genes are located has not been identified in hexaploid oat it is impossible to delete the appropriate chromosome to induce pairing between the alien chromosome and its homoeologous partners in A. sativa. However, the use of a genotype of the diploid species A. longiglumis (Cw 57) which has been shown to suppress the activity of the gene or genes controlling regular bivalent pairing in A. sativa (RAJHATHY & THOMAS, 1972), offers a possible approach to gene transfer in Avena. The procedure for the production of such a genotypically induced gene transfer in Avena is described in this paper.

MATERIALS AND METHODS

The original source of the mildew resistance was a genotype of the tetraploid wild oat A. barbata collected in Algeria. The chromosome addition line involving the A. barbata chromosome on which the gene conferring mildew resistance was located, added to the complement of the cultivar Manod, yielded susceptible and resistant progeny. The susceptible progenies were euploid and the resistant plants always included the A. barbata chromosome (THOMAS et al., 1975). A mildew resistant plant which included only the short arm of the A. barbata chromosome was also isolated. The ditelosomic line was used in the present study.

The amphiploid between the A. longiglumis genotype Cw 57 and the A. sativa cultivar Pendek, produced by Dr T. Rajhathy, Ottawa Research Station, Canada was used as the genotype to induce pairing between homoeologous chromosomes. The cultivar Sun II was used in the backcrossing programme.

For cytological studies immature heads were fixed in Carnoys fixative. Anthers were stained in alcoholic hydrochloric acid-carmine (SNOW, 1963) and squashed in 45% acetic acid.

Mildew reaction. All mildew reaction tests were undertaken using race 3 of Erysiphe graminis f.sp. avenae. Seedlings were dusted with spores from a spreader plant at the two leaf stage and scored for mildew infection 7-14 days after inoculation.

Crossing scheme

i A. sativa x A. longiglumis 8x amphiploid x Ditelocentric addition line (Sun II plus I cv. Pendek cw 51

I pair of telocentrics for short arm of A. barbata chromosome)

A. sativa X F, hybrid (2n = 49 + telocentric short arm) cv. Sun II

BC, hybrid (mildew resistant)

x Sun II

BC, Av 1860

In the crossing scheme the Cw 57 x Pendek amphiploid is used since it produces reasonably fertile hybrids when crossed with A. sativa genotypes. The ditelosomic addition line was used as the mildew resistant genotype since it provides a convenient

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cytological marker. The BC, hybrid used to obtain the BC, plant was resistant to mildew.

RESULTS

No cytological observations were made on the backcross hybrids. The single mildew resistant BC, plant (Av 1860) yielded 10 resistant and 72 susceptible seedlings when selfed. One of the resistant F, seedlings had 43 chromosomes and the telocentric was absent. When this particular plant was selfed all the progeny were resistant to mildew and the majority of the progeny had the euploid number of chromosomes, 2n = 42. The resistant euploid plants had regular meiotic behaviour and formed 21 bivalents at metaphase I.

The F i hybrid Av 1860 x Sun II also regularly formed 2 1 bivalents at metaphase I. Therefore all the evidence indicates that the gene conferring mildew resistance had been transferred into the A. sativa genome as a result of a crossover between the telocentric for the short arm of the A. barb&a chromosome and a chromosome of A. sativa, probably a member of its homoeologous group. Confirmation of the recombination was obtained in the hybrid Av 1860 x ditelosomic addition line where the telocentric was found to be paired in a heteromorphic trivalent. The trivalent was either of the chain or frying pan type, in the latter the telocentric always formed the handle of the configuration. The telocentric was paired in 27 out of the 51 cells analysed while the telocentric is only rarely paired in the monotelocentric addition line (THOMAS et al., 1975).

The F, population from the hybrid Av 1860 x Sun II yielded 38 resistant and 15 susceptible seedlings conforming to the 3 resistant : 1 susceptible ratio expected for a single locus segregation. This regular inheritance of mildew resistance suggests that the transmission of the gametes including the recombinant chromosome was normal.

Av 1860 was crossed with the 15 available monosomics of Sun II in an attempt to identify the chromosome of A. sativa involved in the recombinant. The F, populations from all the 15 monosomic F, hybrids did not deviate significantly from the expected 3 resistant : 1 susceptible ratio and it can be concluded that none of the 15 chromosomes represented by the monosomic lines is involved in the recombinant chromosome (Table 1).

Four further possible recombinants have been isolated, but all have a chromosome number in excess of the euploid number. In these lines the mildew gene could have been transferred onto a chromosome of Cw 57, which as a direct result of selection for mildew resistance could have been established as chromosome addition lines. The

relevant crosses have been made to test whether these lines also involve recombinants between the alien chromosome and their corresponding chromosome in A. saliva.

DISCUSSION

The regular meiotic behaviour of the resistant euploid segregate from Av 1860 and the absence of the telocentric for the short arm of the A. barbata chromosome clearly indicate that a segment of the telocentric has been transferred onto a chromosome of A. sativa. Since THOMAS et al. (1975) were unable to isolate a single natural recombinant

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HUGH THOMAS, W. POWELL AND T. AUNG

Table 1. The segregation of resistant and susceptible seedlings in the F, populations of 15 monosomic lines of A. sativa x Av 1860, the genotype including the transfer.

Monosomic line Resistant Susceptible Chi-square (3 : 1) Probability

1 36 8 1.09 0.1-0.3 2 33 9 0.286 0.5-0.7 3 40 14 0.025 0.7-0.9 4 40 10 0.667 0.3-0.5 5 40 12 0.103 0.7-0.9 6 43 17 0.356 0.5-0.7 7 29 14 1.74 0.1-0.3 8 38 12 0.027 0.7-0.9 9 51 15 0.182 0.5-0.7

10 70 17 1.38 0.1-0.3 11 46 16 0.022 0.7-0.9 12 48 12 0.80 0.3-0.5 13 49 14 0.259 0.5-0.7 14 49 15 0.25 0.550.7 15 39 11 0.24 0.5-0.7

involving the gene for mildew resistance from selfed progeny of the monosomic addition line, the recovery of the Av 1860 segregate in the present study can be attributed to the influence of the A. longiglumis genotype. The Cw 57 genotype increases pairing between homoeologous chromosomes in hybrids with A. sativa (RAJHATHY & THOMAS, 1972). The As genome of A. barbata is assumed to be related to the A genome of A. sativa (NISHIYAMA, 1929; THOMAS & JONES, 1964) since the chromosomes pair in the hybrid between the species. This is also the case in the diploid species with the As genome (KIHARA & NISHIYAMA, 1932; MARSHALL & MYERS, 1961). However, in spite of the observed pairing between the As and A genome chromosomes, recombination of the genes of the diploid and tetraploid species and A. sativa was not obtained (THOMAS et al., 1975; SHARMA, 1975; SHARMA & FORSBERG, 1974). In all associations involving As and A genome chromosomes, the chiasmata are always terminal and this could account for the restricted recombinat’ion observed in the hybrid derivative. The low chiasma frequency reported in hybrids between species of lower ploidy and A. sativa is probably a reflection of only partial homology between the As and A genomes (RAJHATHY & THOMAS, 1974). In such a situation the ability of the A. longiglumis genome to suppress the genetic mechanism controlling bivalent pairing and increase the frequency of homoeologous association would also increase the potential for pairing between the A. barbatachromosome and its corresponding chromosomes in A. sativa and the isolation of recombinant genotypes.

Confirmation of the recombination between the A. barbata telocentric and an A. sativa chromosome was provided by the presence of a heteromorphic trivalent in the hybrid between Av 1860 and the A. barbata ditelosomic addition line. The high frequency with which the A. barbata telocentric paired with the recombined chromo- some shows that a substantial segment of the A. barbata chromosome was involved in the crossover. The heteromorphic chain trivalent demonstrates the terminal position of the A. barbata segment. The occurrence of the frying pan type trivalent also shows that the proximal segment of the A. sativa chromosome arm involved in the crossover was

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sufficiently large for a chiasma to be formed. The segment of the A. barbata chromosome incorporated in the recombinant

chromosome did not have any deleterious effect on the transmission of gametes which included the recombinant chromosome. This is not surprising if the transfer involved homoeologous chromosomes since the gene sequence of the A. barbata chromosome would be similar to the segment of the A. sativa chromosome replaced, as shown by the specific substituting ability of the A. barbata chromosome (THOMAS & BHATTI, 1975).

The use of the Cw 57 genotype does increase the prospects for the exploitation of desirable variation in related diploid and tetraploid weed species in the improvement of the oat crop. Transfers based on recombination between homoeologous chromosomes introduce greater specificity than procedures using irradiation induced transfers where the exchanges are random. A further refinement of this technique would be to use substitution lines involving the alien chromosome as the donor parent. In the hybrid with Cw 57 amphiploid the alien chromosome and its corresponding A. sativa chromo- some would both be in the monosomic condition. The removal of the tendency for preferential pairing should increase synapsis between the two related monosomic chromosomes and increase the number of recombinants isolated.

REFERENCES

AUNG, T., H. THOMAS&I. T. JONES, 1977. The transfer of the gene for mildew resistance from Avena barbata (4 x ) into the cultivated oat A. sativa by an induced translocation. Euphytica 26: 623-632.

GAUTHIER, F. M. & R. C. MCGINNIS, 1968. The meiotic behaviour of a nullihaploid plant in Avena sativa L. Can. J. Genet. Cytol. 10: 186-189.

KIHARA, H. &I. NISHIYAMA, 1932. The genetics and cytology of certain cereals. III. Different compatibility in reciprocal crosses of Avena with reference to tetraploid hybrids between hexaploid and diploid species. Jap. J. Bot. 6: 245-305.

MARSHALL, H. G. & W. M. MYERS, 1961. A cytogenetic study of certain interspecific Avena hybrids and the inheritance of resistance in diploid and tetraploid varieties to races of crown rust. Crop Sci. 1: 29-34.

NISHIYAMA, I., 1929. Thegenetics andcytologyofcertaincereals. 1. Morphological and cytological studies on triploid, pentaploid and hexaploid Avena hybrids. Jap. J. Genet. 5: 148

RAJHATHY, T. & H. THOMAS, 1972. Genetic control of chromosome pairing in hexaploid oats. Nature New Biol. 239: 217-219.

RAJHATHY, T. & H. THOMAS, 1974. Cytogenetics of Oats (Avena L.) Misc. Publications of Genet. Sot. Canada No. 2. The Genetics Society of Canada, Ottawa, 91 pp.

RILEY, R. & V. CHAPMAN, 1958. Genetic control of cytologically diploid behaviour in hexaploid wheat. Nature 182: 713-715.

RILEY, R., V. CHAPMAN & R. JOHNSON, 1968. The incorporation of alien disease resistance in wheat by genetic interference with the regulation of meiotic chromosome synapsis. Genet. Res. 12: 199-219.

SEARS, E. R., 1977. Analysis of wheat-Agropyron recombinant chromosomes. In: Interspecific hybridi- zation in plant breeding, pp. 63-73, Madrid, 1977. Proc. of 8th Congress of Eucarpia.

SHARMA, D. C., 1975. Chromosome pairing problems in interploidy transfer of leaf rust resistance in oats. Euphytica 24: 503-510.

SHARMA, D. C. & R. A. FORSBERG, 1974. Alien chromosome substitution: a cause of instability for leaf rust resistance in oats. Crop Sci. 14: 533-536.

SNOW, R., 1963. Alcoholic hydrochloric acid-carmine as a strain for chromosomes in squash preparations. Stain Technol. 38 : 9-l 3.

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THOMAS, H., J. M. LEGGETT & I. T. JONES, 1975. The addition of a pair of chromosomes of the wild oat A vena barbata (2n = 28) to the cultivated oat A. sativa L. (2n = 42). Euphytica 24: 7 17-724.

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