The proportion of hybrids in seed from a seed orchard composed of two larch species (L europaea and L leptolepis)

Original article The proportion of hybrids in seed from a seed orchard composed of two larch species (L europaea and L leptolepis) M Hacker, F Bergmann Abteilung für Forstgenetik und Forstpflanzenzüchtung, Georg-August-Universität, D-3400 Göttingen, Germany (Received 21 January 1991; accepted 17 June 1991) Summary — The proportion of hybrids in the seed produced in an orchard containing one European larch (Larix europaea) clone used as maternal tree and 271 Japanese larch (Larix lepiolepis) clones used as pollinators was determined by isoenzymatic methods. On the basis of the 2 enzyme systems, Shikimate dehydrogenase (SKDH) and NADH dehydrogenase (NDH), the 2 larch species could be unequivocally distinguished in this case and therefore the proportion of hybrids and selfings (individual and clonal) exactly determined. The genetic control of these enzyme systems was analyzed by means of offspring of controlled (including reciprocal) crossings between one European and one Japanese larch. Five samples of different crop years of the hybrid seeds were examined. The proportion of full seeds ranged from 22-60% and the proportion of hybrids in full seed from 68.684.6%, thus showing that this method of producing hybrids is very efficient. A further advantage of the use of 2 enzyme systems is the easier detection of minimum amounts of contamination, which in this case ranged from 1.0-6.7%. The possible influence of the parental genotypes on the genetic structure of hybrids and possible explanations for the origin of the contamination are discussed. larch / hybrid / seed orchard / contamination Résumé — Taux d’hybridation dans la descendance d’un verger à graines composé de deux espèces de mélèze (Larix europaea et L leptolepis). Le taux d’hybridation dans les lots de graines produits dans un verger contenant un clone de mélèze d’Europe (Larix europaea) utilisé comme arbre mère et 271 clones de mélèzes du Japon utilisés comme pollinisateurs a été déterminé par voie isoenzymatique. À l’aide de 2 systèmes isoenzymatiques : shikimate déshydrogénase (SKDH) et NADH déshydrogénase (NDH), on a été capable de distinguer clairement les 2 espèces et ainsi de déterminer exactement la fréquence d’hybrides et d’autofécondations. Le contrôle génétique de ces systèmes d’enzymes a été analysé à l’aide des descendants de croisements contrôlés (y compris les réciproques) entre un mélèze d’Europe et un mélèze du Japon. Cinq échantillons de graines hybrides récoltés différentes années ont été examinés. Le taux de graines pleines s’élève de 22% à 60% et la proportion d’hybrides dans les graines pleines de 68,6% à 84,6%, montrant ainsi que cette méthode de production de graines hybrides est très efficace. L’utilisation de 2 systèmes d’enzymes facilite, en outre, la détection de quantités minimes de contamination (1,0-6,7%) dans notre cas. L’influence exercée éventuellement par les génotypes parentaux sur la structure génétique des hybrides, et des hypothèses sur l’origine de cette pollution pollinique sont discutées. mélèze / hybride / isoenzyme / verger à graines / contamination INTRODUCTION Forest tree breeding is principally concerned with the improvement of growth, wood quality and vitality in tree species of economic interest. In order to realize such breeding aims as rapidly as possible, eg in one generation, the artificial generation of hybrids between 2 compatible species is performed, because possible combinations of desirable traits of both species are expected in their progeny. Our example of such interspecific hybrids, which have already shown hybrid vigour for many trait combinations, is Larix x eurolepis (Den- gler, 1941; Hering et al, 1989), resulting from crosses between European larch (Larix decidua Miller or L europaea DC) and Japanese larch (Larix kaempferi Sargent or L leptolepis Gord) (for review and litera- compilation, see Langner, 1952, 1971; Kleinschmit, 1988; Paques, 1989). ture The hybrid seed collection designated LOLA 1 (Larix x eurolepis) investigated in this study is the result of part of a breeding program with the aim of developing an inexpensive method for mass-producing hybrid seed material. For this purpose the seed production is based on windpollination of single European larch clones, which are selected according to their general combining ability with Japanese larch, by many Japanese larch clones: one grafted clone of European larch is planted in a Japanese larch stand or seed orchard; the European larch clone is used as maternal tree, the only tree from which seed is harvested, whereas the Japanese larch clones serve as pollen parents. The seed material collected from the European clone is therefore assumed to be a mixture of hybrids (outcrossed fullsibs or half-sibs) and selfings (individual and clonal). This method of producing hy- brid seed was expected to provide high proportions of hybrids (Stern, 1966). Further aims of this breeding strategy are: 1) to obtain a certain uniformity and repeatability of different hybrid seed collections by using one European larch clone as maternal tree; and 2) to obtain the greatest possible genetic variability in the hybrid seed collections by using many Japanese larch clones as pollinators (in contrast to seed orchards with few - sometimes only 2 - clones). The results are genetically different hybrid seed collections, which are the basis for multisite ex- periments to study the stability of hybrid vigour over a range of environments, as suggested by Paques (1989). Since 1971 about 100 different hybrid seed collections have been tested in 30 field experiments distributed over Germany, Austria and France (Langner, in preparation). But hybrid breeding programs based upon wind pollination make no sense, however, unless one can be sure that hybrids will actually be obtained. To our knowledge there exist only a few studies which have identified the proportion of hybrids (Adams and Coutinho, 1977; Joly and Adams, 1983; Bergmann and Ruetz, 1987), but some of these estimates are very uncertain. The determination of the proportion of hybrids should be possible by the use of isoenzyme gene markers similar to the method described by Bergmann and Ruetz (1987). Since the maternal trees are one clone and thus always have the same genotype (in contrast to seed orchards where seed is collected from different clones), there was a good chance to detect isoenzyme markers with sufficient differences between the parent species to identify the hybrids. The determination of the proportion of hybrids was carried out in 4 steps. Species distinction by enzyme systems MATERIALS AND METHODS Preliminary investigations of enzyme sys- Species distinction by enzyme systems tems in order to detect suitable isoenzyme markers, ie enzymes of which European and Japanese larches have distinct isoenzyme variants/alleles. One possible enzyme system was already known: Shikimate dehydrogenase (SKDH). However, this system had onl (...truncated)