A simple and rapid method for estimating representation of species in spruce seedlots using chloroplast DNA restriction fragment length polymorphism

Spruce seedlots containing species mixes and hybrids of Sitka spruce (Picea sitchensis (Bong.) Can.) and interior spruce (Picea glauca (Moench) Voss/P. engelmanni (Parry)) produce seedlings of unacceptable stocktype under operational nursery growing regimes in British Columbia. We have investigated the utility of chloroplast DNA (cpDNA) restriction fragment length polymorphisms for identification of the species composition of these seedlots. A BamHI library of Sitka spruce cpDNA was constructed in pUC8. Two clones were selected by hybridization with a 10.5kb BamHI fragment of white spruce cpDNA which is unique to interior spruce. One of these (pSS4) containing a 4.3kb BamHI fragment was tested in screening of pure and mixed seedlots of Sitka and interior spruce. The results show that this probe can be used to screen total DNA samples to reliably identify and quantify the cpDNA composition of two week old germinants using a sample size of 0.5 g and allows less than 5% species contamination to be detected.