## Doctoral Dissertations

Winter 1991

Dissertation

Biochemistry

#### Degree Name

Doctor of Philosophy

Anita S Klein

#### Abstract

Restriction fragment length polymorphism (RFLP) variation was examined among samples from the entire range of red spruce (Picea rubens Sarg.), the eastern complex of black spruce (Picea mariana (Mill.) B.S.P.), control-cross red-on-black hybrids, and natural populations of red and black spruce. Within-species and population variation was examined. In addition an accurate species index capable of identifying red spruce, black spruce and hybridization between the two species was developed.

The nuclear rDNA repeat unit size in Picea ranged from a minimum of 32 kbp to greater than 40 kbp, two to three fold larger than the typical angiosperm rDNA unit. At a size greater than 32 kbp and a concentration averaging 1.2-1.3 $\times$ 10$\sp4$ copies per pg genomic DNA, the rDNA repeat constitutes approximately 4% of the spruce genome.

The rDNA repeat units were found to be polymorphic within an individual genome with up to five distinct rDNA repeat unit types (alleles) evident. The RFLPs observed in the rDNA repeat were not species specific; however, noticeable trends in internal allelic frequencies were noticed which were useful for between-species differentiation. One marker (EMW 4.35) displayed a significant relationship with geographic origins and habitat suggesting that the observed between-species variation for this marker may be due to selection rather than the result of a true species difference.

Variation in the nuclear rDNA repeat could not accurately differentiate hybrids from black spruce. Additional markers were required to identify hybrids.

RFLPs were identified for the organelle genomes of red spruce, and black spruce. The organelle inheritance pattern was deduced using controlled-cross hybrids. Organelle markers were combined with allelic data from the nuclear rDNA repeat to derive a simple three character index capable of identifying red spruce, black spruce and hybridization between the two species.

Significant gene flow was observed between red and black spruce populations located at Head Harbor, Isle au Haut, Maine but not on Mount Washington or Mount Lafayette. These findings suggest that hybridization and introgression between red and black spruce is influenced by not only proximity of the two species but also by habitat type.

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