Daniele Rosellini1, Franco Lorenzetti1 and Edwin T. Bingham2

1Istituto di Miglioramento Genetico Vegetale, Università degli Studi di Perugia, Borgo XX giugno 74, 06100 Perugia, Italy. E-mail roselli@krenet.it

2Department of Agronomy, University of Wisconsin-Madison, 1575 Linden Dr., Madison, WI, USA 53706

Seed/ovule ratio per unit area in the field is about 8% in alfalfa (1). Among the possible mechanisms of low seed set, embryo abortion has been demonstrated. However, it is not known whether pre-fertilization ovule sterility plays a role. Only a single factor female sterility trait has been described to date (2).

We investigated the inheritance of an ovule sterility trait associated with callose (b1,3 and b1,4 glucan) deposition in mature ovules and studied its occurence in six alfalfa populations. All the plants used were grown in pots in a greenhouse in Madison, Wisconsin, USA, from November 1996 to July 1997 under continuous light. Florets were sampled at flowering, fixed in 70% ethanol:acetic acid 3:1 for at least 24 hours. The pistils were then dissected, cleared in 8N NaOH for 5 hours, stained overnight in 0.1% aniline blue in 0.1M K3PO4, mounted in the staining solution and gently squashed under a coverslip. Fluorescence microscopy using UV light was then used for evidencing callose.

Ovule sterility was found to be associated with callose deposition in B17, a plant with low fertility from the alfalfa cv Blazer XL. The site of callose deposition, which began during embryo sac development and affected 81% of the ovules in mature florets, at random positions in the ovary, appeared to be the embryo sac wall or the integumentary tapetum. B17 was crossed with P13, a Peruvian plant with 5% callosized ovules, to generate reciprocal F1 populations, and an F1 plant (91% callosized ovules) was used to obtain the backcross populations. B17 was also crossed to unrelated, highly fertile plants. S1 progenies from B17 and P13 were also studied.

The fertile ovules of B17 transmitted the ovule sterility trait to the progenies, thereby demonstrating a sporophytic genetic control. All the progeny populations displayed continuous variation for the percentage of sterile ovules, indicating that more than one gene is involved. Reduced transmission of the sterility trait through the pollen is hypothesized to explain the difference between reciprocal crosses. Six progeny plants showing 100% callosized ovules proved to be female sterile. Narrow-sense heritability estimated by offspring-midparent regression was 0.85.

Together, our results demonstrate a quantitative sporophytic additive inheritance of ovule sterility.

The percentage of sterile ovules per plant was determined on 22 to 25 plants from each of four North American and an Italian cultivar, and a tetraploid M. falcata population. On average, 4 to 24% sterile ovules per plant were found in these populations, and plants with up to 75% were observed. Ovule sterility was negatively correlated with the number of seeds per pod with both hand-crossing and selfing. This suggests that seed yield of cultivars could be adversely affected by ovule sterility, and that assessing this trait in base genotypes for cultivar development could help to obtain cultivars with better seed yield.

(1) Lorenzetti F (1993) Achieving potential herbage seed yield in species of temperate regions. Proc XVII International Grassland Congress, Lincoln, New Zealand, 15-16 Feb. pp 1621-1628.

(2) Bingham ET and Hawkins-Pfeiffer J (1984) Female sterility in alfalfa due to a recessive trait retarding integument development. J Hered 75:231-233.

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