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6. DO ANY SPECIES BOUNDARIES....
6.3. RESULTS
6.3.1. Polymorphism between parents of intra-and interspecific crosses
Whether parents originated from the same or different species did not influence the level
of polymorphism that was rather correlated with breeding history of the crossed individuals.
For the 17 enzymes analysed, a total of 34 loci were identified in all parental plants. Among
them 19 loci (55%) were polymorphic between parents of all crosses (Table 6.3). At the rest
of loci but Per2 the same alleles were observed in all analysed parental plants. Polymorphism
was observed as changes in mobility for the majority of loci. The only exception was
the null allele at Est1 in HU5. The number of alleles did not differ much between parents of interspecific
cross, HU5 x BO2 (18%) and both intraspecific crosses (12%). Likewise, DNA differences
were comparable at the intra- and interspecific level. The genetic identity estimated
from RAPD and ISJ markers was only slightly lower for the combination HU5 x BO2 (I=0.754)
than for L. multiflorum (I=0.870) and L. perenne (I=0.842) combinations. The only exception
was the interspecific cross, BR3 x NZ15, whose parents were highly diverse having different
enzymatic alleles at 16 enzymatic loci (37%) and DNA based genetic similarity as low
as I=0.588. Such great differences between BR3 and NZ15 parents were also confirmed by
analyses of other DNA markers including SSR, AFLP, SSAP and bacteria derived-markers,
B-SAP (Figure 6.1). Of the 147 different kinds of primers or primer combinations 43 (29%)
generated various level of polymorphism. They amplified 1018 fragments in total, and among
them 261 were polymorphic (25%) between parents. Another 223 fragments were not polymorphic
between parents; however parental lines should have been heterozygous for them
because these fragments segregated in the F 2
population. Because of high polymorphism
between parents this population was further used for genetic mapping studies.
With regard to enzymatic loci fast alleles were equally distributed between parents of
interspecific crosses (Table 6.4A-B) but they were associated in one of parents in intraspecific
crosses (Table 6.4C-D). Heterozygotes usually had two bands corresponding to parental
bands suggesting that active enzymes are monomers. However, it should be noticed that