Genetic validation and characterization of RAPD markers differentiating black and red spruces: molecular certification of spruce trees and hybrids

Abstract

Picea mariana (black spruce) and P. rubens (red spruce) are closely related species which are difficult to differentiate morphologically. RAPD markers differentiating black and red spruces have been previously identified. In the present study, genetic validity of these markers was determined using samples representing range–wide provenances. Their applicability for certifying genetic identity of individual black, red trees and their hybrids from several sympatric and allopatric locations was demonstrated. These diagnostic fragments of both red and black spruce were present at a frequency of over 0.95 in allopatric provenances, but at a lower frequency in some sympatric provenances (0.43–1.00). Natural populations of red spruce exhibiting typical red spruce phenotype contained black spruce diagnostic RAPD fragments and black spruces growing in bogs with typical bog black spruce morphology, contained red spruce-specific RAPD markers. Some major RAPD markers were cloned and sequenced. The results reveal an extremely high degree of identity between the random primer and the primer binding sites on the genome. Amplification of black and red spruce genomic DNA with designed primers flanking the species-diagnostic RAPD markers indicates that most of RAPD markers used to differentiate black spruce from red spruce are not species specific since these sequences were detected in several spruce species using a more sensitive detection method.