New matchmaking between details out-of hereditary (P
Metropolitan areas away from Platanthera chlorantha (PS1 and you will PS2, PB1–PB4, circles) and you will Cephalanthera rubra (CK1 and you may CK2, CB1–CB7, triangles) communities inside north-east Poland.
Study town and you may sampling
We examined six P. chlorantha and 9 C. rubra communities inside northern-eastern Poland (Bialowieza and Knyszynska Primeval Forest, Szeszupa river area) for the pure, semi-natural and you can anthropogenic groups of federal and you will surroundings parks, supplies and you may protected elements, such as for example Natura 2000 sites ( Fig. 1). While they are based in secure section, of several occur on the train embankments, with each other paths and you may paths in the forest or even in clearings.
The new sampling process depended to your society size. Leaf trials away from nearly all ramets contained in this communities of each varieties have been removed (except people PS2; Dining table 1); no samples had been taken from broken or very young anyone. 100 and you can ninety-eight samples of P. chlorantha and you will 95 products off C. rubra was in fact amassed. Leaf tissues is actually maintained frost up to it can be held during the ?80 °C, dating app for Sapiosexual pending allozyme research. All collected samples were utilized for allozyme analysis.
N, population size; NS, number of samples analysed; NG/NW, number of generative ramets/number of vegetative ramets; PPOL, percentage of polymorphic loci; A, mean number of alleles per locus; HO, observed heterozygosity; HE, expected heterozygosity; FWas, inbreeding coefficient; G, number of genotypes, G/NS, clonal diversity; GU, number of unique genotypes; GU %, percentage of unique genotypes (Fischer’s exact test: P < 0.05, **P < 0.01, ***P < 0.001); #, sum of parameters.
N, population size; NS, number of samples analysed; NG/NW, number of generative ramets/number of vegetative ramets; PPOL, percentage of polymorphic loci; A, mean number of alleles per locus; HO, observed heterozygosity; HE, expected heterozygosity; FIs, inbreeding coefficient; G, number of genotypes, G/NS, clonal diversity; GU, number of unique genotypes; GU %, percentage of unique genotypes (Fischer’s exact test: P < 0.05, **P < 0.01, ***P < 0.001); #, sum of parameters.
Allozyme polymorphism
Homogenates had been prepared by grinding this new actually leaves in a boundary having 2-mercaptoethanol (1%, v/v). Electrophoresis is actually achieved to your 10% starch ties in and you will Titan III cellulose acetate dishes (Helena Laboratories, Beaumont, Texas, USA) after the practical electrophoretic procedures. Fifteen loci (Adh, Gdh, Got-step one, Got-2, Idh-step 1, Idh-2, Mdh-step one, Mdh-2, Me personally, Pgi, Pgm, 6Pgd, Skd, Sod, Tpi) during the P. chlorantha and you will 16 loci into the C. rubra (Adh, Got-step one, Got-dos, Gdh, Idh-step one, Idh-2, Mdh-step one, Mdh-dos, Myself, 6Pgd, Pgi, Pgm, Skd, Sod, Tpi-1, Tpi-2) had been investigated. A few electrode/serum buffer possibilities were used to answer enzyme options: GDH and you can Had (10% lithium-borate lateral starch serum in the pH 8.2/8.3) and you may MDH, SKD and you can TPI (10% histidine-citrate shield on pH 7.0/eight.0). Enzyme interest staining accompanied Soltis Soltis ( 1989). One other chemical systems (ADH, IDH, Me personally, 6PGD, PGI, PGM, SOD) was in fact screened using Titan III cellulose acetate plates, that have been solved playing with Tris-glycine buffer at the pH 8.6 and Tris-citrate barrier in the pH eight.6 (Richardson, Adams Baverstock, 1986). The newest enzyme staining solutions were predicated on Soltis Soltis ( 1989) and you can Richardson ainsi que al. ( 1986), which have adjustment.
Statistical analysis
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 (PPOL), number of alleles per locus (A), genetic diversity (i.e. observed HO and expected heterozygosity HE) and inbreeding coefficient (FIs). 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 (GU) and the probability that the next ramet sampled would be a different genotype (G/NS; where NS is the number of ramets sampled). POL, A, HO and FAre) and population size were tested with Spearman’s pairwise rank correlations (StatSoft, 1995).
