Species-specific nuclear and chloroplast single nucleotide polymorphisms to distinguish Picea glauca, P-mariana and P-rubens

Picea rubens (red spruce) and P. mariana (black spruce) are closely related species which are difficult to differentiate morphologically. They are sym patric with P. glauca (white spruce) in the northern portion of their range s. In order to identify potential interspecific polymorphisms, the chloropl ast trnK intron and rpl33-psaJ-trnP region were sequenced, and the nuclear- encoded ITS region of the rDNA repeat was partially sequenced. Thirteen chl oroplast and 12 nuclear candidate interspecific single nucleotide polymorph isms (SNPs) were identified. The species-specificity of several SNPs was de termined by surveying DNAs amplified from trees representing range-wide pro venance tests; these included 46 red spruce from 11 provenances, 84 black s pruce from 30 provenances and 90 white spruce from 32 provenances. Two SNPs (1 chloroplast and 1 nuclear), which distinguish black spruce from red and white spruce, were consistent among 96-100% of the trees surveyed. Five SN Ps (4 chloroplast and 1 nuclear), which distinguish white spruce from red a nd black spruce, were consistent among 100% of surveyed trees. These specie s-specific SNPs were used to identify anonymous spruce samples in a blind t est, and their utility for small amounts of tissue, as little as single nee dles, was demonstrated. Scoring these SNPs is much less labor intensive tha n previous molecular methods for taxa differentiation (restriction fragment length polymorphisms or random amplified polymorphic DNAs), therefore they can be applied to large population studies.