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Article

An Economical Approach to Distinguish Genetically Needles of Limber from Whitebark Pine

1
Department of Plant Sciences and Plant Pathology, Montana State University, Bozeman, MT 59717, USA
2
Department of Ecology, Montana State University, Bozeman, MT 59717, USA
3
USDA Forest Service Rocky Mountain Research Station, Missoula Fire Sciences Laboratory, 5775 U.S. Highway 10, Missoula, MT 59808, USA
4
Inventory and Monitoring Division, Greater Yellowstone Network, National Park Service, 2327 University Way Suite 2, Bozeman, MT 59715, USA
*
Author to whom correspondence should be addressed.
Forests 2019, 10(12), 1060; https://doi.org/10.3390/f10121060
Received: 7 November 2019 / Accepted: 20 November 2019 / Published: 22 November 2019
(This article belongs to the Special Issue Ecology and Restoration of Whitebark Pine)
Whitebark pine is difficult to distinguish from limber pine when seed cones are not present. This is often the case because of young stand age, growth at environmental extremes, or harvesting by vertebrate species. Developing an economical genetic identification tool that distinguishes non-cone-bearing limber from whitebark pine, therefore, could aid many kinds of research on these species. Phylogenetic studies involving limber and whitebark pine suggest that chloroplast DNA sequences differ between these species. We therefore wanted to identify chloroplast loci that could differentiate limber from whitebark pine trees by taking an economical approach involving restriction-site analysis. We generated chloroplast DNA barcode sequences sampled from limber and whitebark pine trees that we identified using attached seed cones. We searched for nucleotide differences associated with restriction endonuclease recognition sites. Our analyses revealed that matK and the psbA-trnH spacer each readily amplified and harbored multiple DNA-sequence differences between limber and whitebark pine. The matK coding sequence of whitebark pine has a BsmAI restriction site not found in limber pine. The psbA-trnH spacer of limber pine has two PsiI restriction sites, neither of which is found in whitebark pine. DNA-sequence and restriction-site analysis of the psbA-trnH spacer from 111 trees showed complete congruence between visually and genetically identified limber (n = 68) and whitebark (n = 43) pine trees. We conclude that restriction site analysis of the chloroplast psbA-trnH spacer and matK involves both minimal technical expertize and research funds. These findings should be of value to foresters interested in species identification and distribution modeling, as well as the analysis of fossil pine pollen, given that gymnosperms transmit chloroplast DNA paternally. View Full-Text
Keywords: chloroplast DNA; genetic identification; Greater Yellowstone Ecosystem; haplotypes; Pinus albicaulis; Pinus flexilis; western North America chloroplast DNA; genetic identification; Greater Yellowstone Ecosystem; haplotypes; Pinus albicaulis; Pinus flexilis; western North America
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    Link: http://www.montana.edu/mlavin/data/Appendix.txt
    Description: Chloroplast DNA sequences of the psbA-trnH spacer region, the matK coding region, and the trnG intron, and the trnL intron region of limber pine (Pinus flexilis), whitebark pine (P. albicaulis) that we generated during this study, along with selected sequences of pine species represented in GenBank. The numbers above the DNA sequences represent the nucleotide site number. The numbers following the species name refer to DNA isolate (Supplemental Table S1) and GenBank accession number. Annotations include locations of restriction endonuclease recognition sites. Wide formatting requires posting of the Appendix to http://www.montana.edu/mlavin/data/Appendix.txt (copy and paste Appendix.txt contents into text editor without text wrapping.)
MDPI and ACS Style

Alongi, F.; Hansen, A.J.; Laufenberg, D.; Keane, R.E.; Legg, K.; Lavin, M. An Economical Approach to Distinguish Genetically Needles of Limber from Whitebark Pine. Forests 2019, 10, 1060. https://doi.org/10.3390/f10121060

AMA Style

Alongi F, Hansen AJ, Laufenberg D, Keane RE, Legg K, Lavin M. An Economical Approach to Distinguish Genetically Needles of Limber from Whitebark Pine. Forests. 2019; 10(12):1060. https://doi.org/10.3390/f10121060

Chicago/Turabian Style

Alongi, Franklin, Andrew J. Hansen, David Laufenberg, Robert E. Keane, Kristin Legg, and Matt Lavin. 2019. "An Economical Approach to Distinguish Genetically Needles of Limber from Whitebark Pine" Forests 10, no. 12: 1060. https://doi.org/10.3390/f10121060

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