Metropolises regarding Platanthera chlorantha (PS1 and you may PS2, PB1–PB4, circles) and you may Cephalanthera rubra (CK1 and you may CK2, CB1–CB7, triangles) populations in northern-east Poland.

Data area and you may testing

I examined half dozen P. chlorantha and 9 C. rubra communities into the northern-east Poland (Bialowieza and you will Knyszynska Primeval Forest, Szeszupa river area) into the absolute, semi-sheer and you will anthropogenic organizations from national and surroundings areas, supplies and protected section, like Natura 2000 internet ( Fig. 1). While he’s situated in secure portion, of numerous occur with the railway embankments, together routes and you will routes when you look at the forest or in clearings.

The latest sampling techniques depended towards inhabitants dimensions. Leaf products from almost all ramets within communities of every variety have been pulled (except people PS2; Desk step 1); zero trials was obtained from damaged or most younger someone. One hundred and you can ninety-seven products from P. chlorantha and you may 95 trials away from C. rubra have been built-up. Leaf tissues try kept on freeze until it could be stored from the ?80 °C, pending allozyme studies. All collected trials were utilized to own allozyme investigation.

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_Are, 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_Are, 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 were prepared by grinding the will leave for the a barrier with 2-mercaptoethanol (1%, v/v). Electrophoresis was carried out towards 10% starch gels and you can Titan III cellulose acetate dishes (Helena Laboratories, Beaumont, Tx, USA) following basic electrophoretic methods. Fifteen loci (Adh, Gdh, Got-1, Got-dos, Idh-step one, Idh-dos, Mdh-step one, Mdh-dos, Me, Pgi, Pgm, 6Pgd, Skd, Sod, Tpi) inside P. chlorantha and you can 16 loci when you look at the C. rubra (Adh, Got-step one, Got-2, Gdh, Idh-1, Idh-dos, Mdh-step 1, Mdh-dos, Me personally, 6Pgd, Pgi, Pgm, Skd, Sod, Tpi-1, Tpi-2) had been investigated. One or two electrode/gel barrier possibilities were utilized Christian dating to respond to chemical expertise: GDH and you can Got (10% lithium-borate lateral starch solution on pH 8.2/8.3) and you will MDH, SKD and TPI (10% histidine-citrate boundary from the pH eight.0/7.0). Chemical pastime staining accompanied Soltis Soltis ( 1989). Additional enzyme options (ADH, IDH, Myself, 6PGD, PGI, PGM, SOD) have been processed using Titan III cellulose acetate plates, that happen to be solved using Tris-glycine boundary on pH 8.6 and you may Tris-citrate shield at pH seven.6 (Richardson, Adams Baverstock, 1986). This new chemical staining remedies have been centered on Soltis Soltis ( 1989) and you can Richardson et al. ( 1986), which have modifications.

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_{Is actually}). 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).