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Premise of the study: Recently released Pinus plastome sequences support
characterization of 15 plastid Simple Sequence Repeat (ptSSR) loci
originally published for P. contorta and P. thunbergii. This allows
selection of loci for single-tube PCR multiplexed genotyping in any
subsection of the genus. Methods: Unique placement of primers and primer
conservation across the genus were investigated, and a set of six loci
were selected for single-tube multiplexing. We compare interspecific
variation between ptSSRs and nucleotide sequences of ycf1 then test
intraspecific variation for ptSSRs using 911 samples in the P. ponderosa
species complex. Results: The ptSSR loci contain mononucleotide and
complex repeats with additional length variation in flanking regions. They
are not located in hypervariable regions and most primers are conserved
across the genus. A single PCR per sample multiplexed for six loci yielded
45 alleles in 911 samples. Discussion: The protocol allows efficient
genotyping of many samples. The ptSSR loci are too variable for Pinus
phylogenies but are useful for the study of genetic structure within and
among populations. The multiplex method could easily be extended to other
plant groups by choosing primers for ptSSR loci in a plastome alignment
for the target group.
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