Chloroplast and mitochondrial molecular tests identify European x Japanese larch hybrids.

Hybrids between European and Japanese larches combine the properties of both parental species (drought resistance, canker resistance, stem straightness) and exhibit a fast growth rate. They are produced in seed orchards, generally by natural pollination. Seeds are collected and used for afforestation as interspecific hybrids. However, there are no convenient tests to assess the interspecific hybrid proportion. In the present study, we developed diagnostic molecular markers suitable for the individual identification of hybrids, whatever their developmental stage. Our strategy involved testing a combination of maternally inherited markers from the mitochondrial genome (mtDNA) and paternally inherited markers from the chloroplast genome (cpDNA). Hybrids were then identified by the presence of a mitochondrial sequence inherited from one parental species and a chloroplast sequence inherited from the other parental species. To achieve this aim, markers discriminating both parental species were first sought. Amplifications of mitochondrial and chloroplast sequences were performed using specific PCR primers. After testing 33 primer pairs in combination with nine restriction enzymes, we detected one mitochondrial marker, f13 which was amplified in Japanese larch and absent in European larch, and one chloroplast marker, ll- TaqI which showed different restriction patterns depending on the species. A restriction fragment of 601 bp was obtained in Japanese larch while two fragments of 120 bp and 481 bp were observed in European larch. These patterns were found in all 197 individuals tested from the two pure species. These markers were then used for the evaluation of the hybrid proportion in a seed lot produced from seed orchards; this was assessed as between 43% and 53% depending on the parental species. The male and female parental species could be determined for each progeny.

Heterozygosity and hybrid performance in larch.

Random Amplified Polymorphic DNAs (RAPD) were used for estimating genetic distances between 12 European larches (Larix decidua) and 12 Japanese larches (L. kaempferi) that were the parents in a factorial mating design. One hundred and eleven fragments were used for establishing genetic distances based on Jaccard's coefficient between parents. Thirteen fragments differentiated the larch species. The genetic distance between individuals of the same species (D J =0.39 in the Japanese larch and 0.45 in the European larch) was lower than the genetic distance between species (D J =0.72). A UPGMA dendrogram based on genetic distances clearly clustered each larch species, confirming the speciation at a molecular level. Correlations between genetic distances of the parents and performances of the hybrid families were established for various quantitative traits. Significant values were found for growth characters and branch insertion angle, which suggested an effect of general heterozygosity level on hybrid traits. These correlations also evolved with tree age: the maximal correlation was noticed on 6-year-old trees for height. The lack of correlation between parental genetic distances and hybrid performances for the other quantitative traits suggested that these characters were controlled by fewer genes. The results of this study show that crosses between genetically distant parents produce hybrids with excellent growth performances; this represents a potential selection criterion of the genitors.