Lendvay et al. (Suppl Mat.) – 1

Article title: Genetic structure in Pinus cembra from the Carpathian Mountains inferred from nuclear and chloroplast microsatellites confirms postglacial range contraction and identifies introduced individuals

Journal name: Tree Genetics & Genomes

Author names: Bertalan Lendvay, Mária Höhn, Sabine Brodbeck, Marcel Mîndrescu, Felix Gugerli

Corresponding author:

Name: Felix Gugerli

Affiliation: WSL Swiss Federal Research Institute, Birmensdorf, Switzerland

E-mail address:

Lendvay et al. (Suppl. Mat.) – 2

Table S1. Basic information on newly developed nuclear microsatellite loci for Pinus cembra, on the basis of Roche 454 sequences (cf. Schoebel et al. 2013). For primer concentrations in multiplex PCR, see Table S2.

Locus / Repeat motif / Primer sequences (5'®3') / Fluorescent dye (on F primer) / Ta [°C] a / Size range b / Allele number b / GenBank accession
Pc-HJM / [TC]10 / F GAAAGGTCACATGTTGCACG / FAM / 58 / 168–188 / 6 / KF692109
R CACATTCTCCAGAGCACTCC
Pc-BUG / [GA]13 / F CTTCAATGCGGCTTCAGAAT / FAM / 58 / 101–133 / 17 / KF692108
R GAGTAGAAGAACAAGTTCTAAGTTTGG
Pc-YAU / [TCA]7 / F TCCTTGCCTCATTTTCATCG / ATTO550 / 58 / 144–162 / 5 / KF692110
R AGCACAGTGAAGGTCACAGC
Pc-28Z / [CA]9G [AC] 5 / F AGTTGACTGTGCCGTCATATTG / FAM / 58 / 288–324 / 8 / KF692111
R CGCATTCCCTAATGCCAAGC
Pc-CQG / [AC]8[AT]5 / F TGTCATGAAGAAATAACCAGGAGA / YakimaYellow / 58 / 169–179 / 6 / KF692107
R TTCCTAGTTTGGTGTGTGGTTG
Pc-SWK / [AT]10 / F TTACCAAGTAAACCCTTTGGTG / ATTO550 / 58 / 236–318 / 46 / KF692112
R TCACTAATCGACATGATTCTTACC

a PCR reactions were multiplexed using Type-it (Qiagen, Hilden, Germany)

b Values relate to data of Carpathian (this article) and Alpine populations (Gugerli & Brodbeck, unpubl. data)


Table S2. Primer characteristics of the three multiplex PCR mixtures used in this study on Pinus cembra. Each forward primer was labeled with a fluorescent dye, end concentrations of the primers were optimized to produce similarly strong PCR amplification in the multiplex reactions

Chloroplast microsatellites
(Vendramin et al. 1996) / Nuclear microsatellites
(Salzer et al. 2009) / Nuclear microsatellites
(454-based; this study) a
Locus / Labeling / Concentration [µM] / Locus / Labeling / Concentration [µM] / Locus / Labeling / Concentration [µM]
Pt630718 / FAM / 0.064 / Pc1b / FAM / 0.20 / Pc-HJM / FAM / 0.075
Pt15169 / FAM / 0.127 / Pc7 / FAM / 0.25 / Pc-BUG / FAM / 0.100
Pt36480 / HEX / 0.212 / Pc18 / NED / 0.20 / Pc-YAU / ATTO550 / 0.100
Pt26081 / HEX / 0.169 / Pc22 / PET / 0.40 / Pc-28Z / FAM / 0.100
Pc23 / VIC / 0.20 / Pc-CQG / YakimaYellow / 0.100
Pc35 / PET / 0.20 / Pc-SWK / ATTO550 / 0.100

a Details on newly developed microsatellites are given in Table S1.


Table S3. Pairwise population differentiation values: FST values for nuclear microsatellites above diagonal and ΦPT values for chloroplast microsatellites below diagonal. Significant p-values (p<0.05) based on 9999 randomization steps are marked with *, highly significant p-values (p<0.001) are marked with ***

Mor / Vel / Ked / Lal / Nea / Neg / Cin / Mun / Gen / Gem
Mor / - / 0.012 / 0.046* / 0.018 / 0.035* / 0.044* / 0.021* / 0.064*** / 0.098*** / 0.044*
Vel / 0.068* / - / 0.054*** / 0.024* / 0.035* / 0.052*** / 0.035* / 0.061*** / 0.099*** / 0.06***
Ked / 0.015 / 0.094* / - / 0.04* / 0.041* / 0.089*** / 0.052* / 0.113*** / 0.157*** / 0.06***
Lal / 0 / 0.071* / 0 / - / 0.014* / 0.038* / 0.033* / 0.066*** / 0.111*** / 0.043*
Nea / 0.077* / 0.037 / 0.078* / 0.047 / - / 0.001 / 0 / 0.051*** / 0.115*** / 0.047***
Neg / 0.034 / 0.083* / 0.046 / 0.03 / 0.023 / - / 0 / 0.048*** / 0.106*** / 0.071***
Cin / 0.122 / 0.097* / 0.176* / 0.105* / 0.135* / 0.187* / - / 0.056* / 0.068* / 0.062*
Mun / 0.214*** / 0.103* / 0.232*** / 0.177*** / 0.081* / 0.185*** / 0.228* / - / 0.041* / 0.093***
Gen / 0.122* / 0.041 / 0.127* / 0.09 / 0.094* / 0.128* / 0.190* / 0.135* / - / 0.133***
Gem / 0.112* / 0.028 / 0.133*** / 0.091* / 0.027 / 0.104* / 0.122* / 0.045 / 0.018 / -

Lendvay et al. (Suppl. Mat.) – 2

a

b

Fig. S1. STRUCTURE (Pritchard et al. 2000) results of the 175 Carpathian Pinus cembra individuals: a Ln Probability of data plotted against number of K clusters and b bar plots showing individual assignment for K=2-5. Specimens appearing in blue color in populations BOR and LAL were identified as introduced.
a

b

Fig. S2. STRUCTURE (Pritchard et al. 2000) results of 624 native and introduced Pinus sibirica, Eastern Alpine and Carpathian Pinus cembra individuals: a Ln Probability of data plotted against number of K clusters and b bar plots showing individual assignment for K=6-8.