Brand new matchmaking ranging from variables of genetic (P

Places from Platanthera chlorantha (PS1 and you can PS2, PB1–PB4, circles) and Cephalanthera rubra (CK1 and you can CK2, CB1–CB7, triangles) populations for the north-eastern Poland.

Data area and you will testing

We investigated half dozen P. chlorantha and you can 9 C. rubra communities within Women’s Choice dating the north-eastern Poland (Bialowieza and Knyszynska Primeval Tree, Szeszupa river area) inside natural, semi-pure and you can anthropogenic teams away from national and you can landscape areas, reserves and safe areas, eg Natura 2000 sites ( Fig. 1). Although he could be situated in safe section, many are present on the railway embankments, collectively channels and you will pathways in the forest or even in clearings.

The latest sampling processes depended with the society size. Leaf examples regarding the majority of ramets within communities of any varieties have been removed (except populace PS2; Table step 1); zero examples was obtained from damaged otherwise really younger some one. One hundred and you will 90-7 samples away from P. chlorantha and you will 95 samples regarding C. rubra have been amassed. Leaf cells is actually kept on freeze up until it could be stored in the ?80 °C, pending allozyme data. All collected samples were used getting allozyme research.

N, population size; N_S, number of samples analysed; N_Grams/N_W, number of generative ramets/number of vegetative ramets; P_POL, percentage of polymorphic loci; A, mean number of alleles per locus; H_O, observed heterozygosity; H_E, expected heterozygosity; F_{Is actually}, inbreeding coefficient; G, number of genotypes, G/N_S, clonal diversity; G_U, number of unique genotypes; G_U %, percentage of unique genotypes (Fischer’s exact test: P < 0.05, **P < 0.01, ***P < 0.001); #, sum of parameters.

N, population size; N_S, number of samples analysed; N_G/N_W, number of generative ramets/number of vegetative ramets; P_POL, percentage of polymorphic loci; A, mean number of alleles per locus; H_O, observed heterozygosity; H_E, expected heterozygosity; F_Is, inbreeding coefficient; G, number of genotypes, G/N_S, clonal diversity; G_U, number of unique genotypes; G_U %, percentage of unique genotypes (Fischer’s exact test: P < 0.05, **P < 0.01, ***P < 0.001); #, sum of parameters.

Allozyme polymorphism

Homogenates have been prepared by milling the latest simply leaves for the a boundary which have 2-mercaptoethanol (1%, v/v). Electrophoresis try accomplished into the ten% starch ties in and Titan III cellulose acetate plates (Helena Labs, Beaumont, Tx, USA) adopting the fundamental electrophoretic steps. Fifteen loci (Adh, Gdh, Got-step 1, Got-2, Idh-step 1, Idh-dos, Mdh-step 1, Mdh-dos, Me, Pgi, Pgm, 6Pgd, Skd, Sod, Tpi) inside P. chlorantha and you will 16 loci when you look at the C. rubra (Adh, Got-step 1, Got-dos, Gdh, Idh-step one, Idh-2, Mdh-1, Mdh-dos, Me personally, 6Pgd, Pgi, Pgm, Skd, Sod, Tpi-1, Tpi-2) was basically investigated. Several electrode/solution barrier solutions were utilized to resolve chemical solutions: GDH and you will Had (10% lithium-borate lateral starch gel from the pH 8.2/8.3) and you can MDH, SKD and you can TPI (10% histidine-citrate buffer from the pH 7.0/seven.0). Enzyme craft staining adopted Soltis Soltis ( 1989). Others enzyme solutions (ADH, IDH, Myself, 6PGD, PGI, PGM, SOD) was processed having fun with Titan III cellulose acetate plates, which were resolved having fun with Tris-glycine buffer from the pH 8.6 and you can Tris-citrate shield from the pH eight.6 (Richardson, Adams Baverstock, 1986). The newest chemical staining formulas was indeed predicated on Soltis Soltis ( 1989) and Richardson mais aussi al. ( 1986), which have changes.

Analytical studies

The data matrix of individuals was analysed using the TFPGA package (Miller, 1997), FSTAT 2.9.3 (Goudet, 2001) and GENEPOP 3.2 (Raymond Rousset, 1995) for calculation of standard measures of allozyme diversity: allelic frequencies, percentage of polymorphic loci (P_POL), number of alleles per locus (A), genetic diversity (i.e. observed H_O and expected heterozygosity H_E) and inbreeding coefficient (F_Try). The occurrence of unique alleles was used to describe population distinctiveness (Slatkin, 1985). Deviations from Hardy–Weinberg expectations were tested for the population by the Markov chain method (GENEPOP).

Parameters of within-population genotypic diversity were also estimated. Three different measures of clonal diversity were used: number of observed genotypes (G), number of genotypes unique to a single population (G_U) and the probability that the next ramet sampled would be a different genotype (G/N_S; where N_S is the number of ramets sampled). _POL, A, H_O and F_{Is actually}) and population size were tested with Spearman’s pairwise rank correlations (StatSoft, 1995).