Russian Journal of Genetics, Vol. 37, No. 2, 2001, pp. 162–167. Translated from Genetika, Vol. 37, No. 2, 2001, pp. 223–229. Original Russian Text Copyright © 2001 by Fedorenko, Savushkin, Olimpienko.
PLANT GENETICS
Genetic Diversity in Natural Populations of Arabidopsis thaliana (L.) Heynh. from Karelia O. M. Fedorenko, A. I. Savushkin, and G. S. Olimpienko Institute of Biology, Karelian Research Center, Russian Academy of Sciences, Petrozavodsk, 185610 Russia; fax: (8142)77-98-10; e-mail:
[email protected] Received June 21, 2000
Abstract—The genetic structure of ten natural populations of Arabidopsis thaliana (L.) Heynh. at eight isozyme loci was studied. The populations were located in the northern part of the species range, 200 km from the north to the south along the Onega Lake coast in Karelia. Considerable genetic diversity (P99% = 43.7, Hobs = 0.003) was revealed that is not typical of populations of self-pollinating plant species. A direct correlation between the proportion of polymorphic loci and geographical latitude was shown (r = 0.68; P < 0.05). It is suggested that a high polymorphism level in Karelian Arabidopsis thaliana (L.) populations increasing from the south to the north is due to extreme environmental conditions in the northern part of the species range. The distribution of genetic diversity within and between populations is typical of self-pollinating species: the larger part of the total diversity resides among populations (GST = 0.583).
Arabidopsis thaliana (L.) Heynh. has long been used as a model species in genetic studies. Most important progress has been achieved in developmental genetics, cell biology, and physiology of A. thaliana. Although A. thaliana is a self-pollinating species, high intraspecific variability of morphological characters, chromosome complements [1], and the time of flowering has been observed [2, 3]. A. thaliana has several advantages over other species making it a convenient model for genetic analysis in higher plants. However, the population genetics of this species is poorly studied. Karelia belongs to the extreme northern part of the species range. In such marginal regions, populations are subjected to high pressure of unfavorable ecological conditions. Two opposite concepts on the features of the population-genetic structure in marginal populations have been advanced. Some authors assume that marginal populations are often monomorphic [4–6]. Levontin, conversely, stressed instability of environmental conditions at the periphery of the species range resulting in “… selection of different genotypes in different periods. It is not, therefore, surprising that gene heterozygosity here is high…” [7, 8]. In Karelia, A. thaliana typically grows on scales with poor soils often in communities with similar plant species. Some populations are very small. The most northern population was found at a latitude of 62°12′; however, it cannot be excluded that some populations occupy even more northern regions. In this paper, we report our results on the population-genetic structure of A. thaliana from the northern part of its range. MATERIALS AND METHODS Seeds of A. thaliana were collected from ten populations occupying in Karelia the Onega Lake coast from
the north to the south. The characteristics of habitats of the populations and their geographical locations are given in Table 1. Seeds were germinated in Petri dishes on agar growth medium under luminescent lamps according to Gichner-Weleminskii. Three-week-old plants were analyzed by electrophoresis. To determine the time of flowering, A. thaliana was transferred from Petri dishes to the soil (soil : sand, 1 : 1). Preparation of leaf samples for electrophoresis, separation, and staining of isozymes were conducted by standard procedures [9]. Electrophoresis was conducted in slabs of polyacryalamide gel in Tris-EDTA-borate buffer, pH 8.3, at U = 200 V for 2.5 h. To study the genetic control of several enzyme loci, crosses between plants of particular genotypes were performed. Populations were analyzed at eight isozyme loci of four enzyme systems: isocitrate dehydrogenase (IDH, E.C. 1.1.1.42), glutamic-oxaloacetic transaminase (GOT, E.C. 2.6.1.1), acid phosphatase (ACP, E.C. 3.1.3.2), and esterase (EST, E.C. 3.1.1*). These enzymes have been well studied in A. thaliana. IDH is a dimer encoded by one locus with two codominant alleles [10, 11]. GOT is controlled by two independent loci. Each locus has two alleles with codominant inheritance [12]. Jacobs and Schwind [13], as well as Abbot and Gomes [14], isolated three activity zones in the acid phosphatase spectrum resulting from allelic segregation at three loci. The genetic control of esterase has not been studied in detail so far. In the multiband pattern, Grover [12] isolated two activity zones and showed that allelic variation was due to variability at two loci (Est-1 and Est-2). We designated loci revealed by histochemical staining in accordance with electrophoretic mobility of their gene products: a protein with the highest mobility towards the anode was denoted by 1; the next protein,
1022-7954/01/3702-0162$25.00 © 2001 MAIK “Nauka /Interperiodica”
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Table 1. Characteristics of the habitats of A. thaliana Karelian populations Population Kondopoga Konchezero Kosalma Tsarevichi Suisar’ Pin’guba Chertov Stul Petrozavodzk Sheltozero Rybreka
Geographic location
Growing site
62°12′ N 34°16′ E 62°08′ N 34°01′ E 62°01′ N 34°07′ E 62°01′ N 34°07′ E 61°58′ N 34°35′ E 61°53′ N 34°33′ E 61°50′ N 34°24′ E 61°48′ N 34°17′ E 61°22′ N 35°23′ E 61°16′ N 35°34′ E
Suburb, along the highway, on a rock near the garden Along the pathway in the village Along the highway, on a rock Along the highway, on a rock near the lake Sand mound at the dirt road Along the highway, on a rock Petrozavodsk suburb, on high steep rock Along the highway, on a rock Along the paved road, on a rock Stone quarry, on the shelf of a high rock
by 2, etc. Alleles were designated by letters according to their electrophoretic mobility as follows: F, fast; S, slow, and I, intermediate. The null allele was denoted by 0. The remaining alleles and rare variants were assigned arbitrary letters. The statistical analysis of the data was carried out by standard procedures used in [15] using the BIOSYS-1 program [16]. To study the genetic subdivision of populations, Nei’s statistics of gene diversity were used [17]. Cross-pollination frequency was estimated from the equation suggested by Li [18]. RESULTS AND DISCUSSION Genetic Analysis of Four Enzyme Loci Genetic analysis of fast and slow isozymes at Idh-1 and Got-1 confirmed the one-locus control of each enzyme with codominant inheritance of two alleles at each of the two loci. Both enzymes were dimeric and characterized by the occurrence of a hybrid band in heterozygotes. The second locus of glutamic-oxaloacetic transaminase (Got-2) encoding a protein with the highest anode mobility was monomorphic in all populations. Staining of acid phosphatase revealed five activity zones; the most anode component was represented by one invariant band in all plants. The genetic analysis of Acp-2 showed that the enzyme was dimeric with codominant inheritance of both alleles. Three bands were observed in heterozygotes. Two single-banded variants were revealed in the most rapidly migrating esterase activity zone (Est-3). In F1 hybrids, both bands were present, and segregation in F2 conformed to the 1 : 2 : 1 ratio. Thus, locus Est-3 encodes a monomer with codominant inheritance of alleles. Allele Frequencies at Eight Enzyme Loci Of eight loci analyzed (Idh-1, Got-1, Acp-2, Acp-3, Est-1, and Est-3), six were polymorphic, and two (Got-2 and Acp-5) were monomorphic and represented by one invariant band in all populations studied. Allele frequencies at polymorphic loci are shown in Table 2. RUSSIAN JOURNAL OF GENETICS
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Allele composition in populations was similar only at locus Got-1: in all populations, the F allele was common. Almost the same trend was observed for locus Est-3, except for two population, Kosalma and Suisar’, in which the F allele was less frequent than the S allele. At other polymorphic loci, most common alleles were different, which is characteristic of self-pollinating species. It should be stressed that the Petrozavodsk population occupied a special place with respect to locus Acp-3. Only in this population, the F allele was the most frequent (0.976). Note that the population from Petrozavodsk had a very low germinating capacity (20.6%). Seed germination in other populations was much better (on average 84.4%). It may due to the unfavorable ecological situation in the city. The population occupied regions along the highway and near industrial plants. All germinated plants were monomorphic at all loci, except for Acp-3. Such plants may be resistant to technogenic factors. Polymorphism for the Time of Flowering Analysis of populations for the time of flowering showed that the Karelian populations were mainly represented by late-flowering (winter) forms. This finding confirmed the observation of Kranz and Kircheim [19] that the latest ecotypes of A. thaliana often grow in the most northern countries. They may have selective advantage over early-flowering forms under the conditions of short and cold summer. Our data indicate that without cold pretreatment of seeds, plants of five populations (Kondopoga, Konchezero, Kosalma, Suisar’, and Petrozavodsk) flowered between 4 and 6 months (120 days and more) after germination; in the Rybreka population, plants began flowering 9 months after germination (270 days and more), and four populations were polymorphic for the flowering time: they were represented both by early-flowering (spring) and lateflowering forms. The proportions of plants (in percents), which began flowering 45, 60, 75, and 90 days after sowing, are given in Table 3. Note for comparison that Cetl [20] considered A. thaliana plants that flow2001
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Table 2. Allele frequencies at polymorphic loci in Karelian populations of A. thaliana Population Locus Tsarevichi
Petrozavodsk
Rybreka
KondoKonchezero Kosalma Sheltozero Pin’guba poga
Suisar’
Chertov Stul
76
103
56
71
16
Idh-1 (N)
78
F
0.891
S
0.109
1.000
1.000
61
0.268 0.732
51
50
0.314 1.000
0.686
1.000
54 0.898
0.239
0.102
0.761
1.000
Got-1 (N)
82
F
0.976
S
0.024
55 1.000
104 1.000
70 0.707
70 1.000
56 1.000
64 1.000
0.293
70
53
17
0.914
0.962
0.086
0.038
1.000
Acp-2 (N)
74
F
0.027
S
0.973
50
54 1.000
1.000
51
51
36
0.755
0.098
0.806
0.245
0.902
0.194
48
57
70
0.035 1.000
16
1.000
1.000
0.965
Acp-3 (N)
100
F
84
59
53
76
0.976
S
0.490
I
0.510
0.024
47 0.043
0.288
0.226
0.339
0.717
51
0.745
72
16
0.078
0.109 0.164
0.139
0.922
0.673
0.677
0.149 1.000
55
0.194
0
0.750 0.250
E
0.373
G
0.064 0.057
0.055
Est-1 (N)
73
F
0.068
S
0.932
50
118
78
86
1.000 1.000
44
95
54
91
0.523 1.000
1.000
0.447
25
0.110 1.000
1.000
0.890
1.000
Est-3 (N)
53
F
0.821
S
0.160
0
0.019
50 1.000
62 1.000
47 0.851
84 0.976
0.149
44
58
54
68
0.432
0.888
0.926
0.382
0.920
0.568
0.112
0.074
0.515
0.080
0.024
ered 28 days after germination to be early-flowering. It is of particular interest that four populations polymorphic for the time of flowering grew in the southern part of the region examined. However, adjacent to the Sheltozero population polymorphic for the time of flowering, the most late-flowering Rybreka population was located. Genetic Diversity The main parameters of genetic diversity of the populations (the average number of alleles per locus (A),
25
0.103
the proportion of polymorphic loci (P) at 95% and 99% criteria, and observed (Hobs) and expected (HÂı) heterozygosities) are presented in Table 4. The average number of alleles per locus varied from 1.1 in Petrozavodsk to 1.9 in the Kosalma, Tsarevichi, and Pin’guba populations. The observed heterozygosity was noted in four populations: in the most northern Kondopoga population and in three polymorphic for the time of flowering populations: Tsarevichi, Pin’guba, and Sheltozero. The mean observed heterozygosity was 0.003. Expected heterozygosity used to measure genetic diversity is more informative for self-pollinating popu-
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Table 3. Polymorphism for the time of flowering in four populations of A. thaliana
(0.6%). Mean cross-pollination frequency in the Karelian populations of A. thaliana was 0.56%.
Number of flowering plants, % Onset of flowering 45 days 60 days 75 days 90 days
Populations containing both early- and late-flowering forms were genetically closer to cross-pollinating species, since the cross-pollination frequency and observed heterozygosity were higher in these populations than in late-flowering forms, in which, in turn, the expected heterozygosity was high. Accordingly, although heterozygotes for the loci studied were not revealed in the late-flowering populations (except for the most northern population), genetic heterogeneity of these populations was rather high. In all likelihood, late-flowering forms as true self-pollinators possess their own mechanisms providing genetic heterogeneity: selection and limited recombination promote the formation of genotypes based on coadaptive gene complexes [26, 27]. Just in the late-flowering populations, a correlation between frequencies of the F allele at locus Idh and the I allele at locus Acp-3 (r = –0.95; P < 0.01) and between frequencies of the F allele at locus Acp-2 and S allele at locus Acp-3 (r = 0.88; P < 0.05) was shown.
Population Tsarevichi Sheltozero Pin’guba Chertov Stul
39st day 37th day 67th day 62nd day
16.2 12.9 0 0
43.2 12.9 0 0
75.7 22.6 6.3 2.5
86.5 45.2 18.8 22.5
lations. As seen from Table 4, the HÂı values were on average higher in late-flowering populations than in populations consisting both of spring and winter forms. An increase in the expected heterozygosity and in the proportion of polymorphic loci in populations from the south to the north, with the highest polymorphism level in the north (62.5%), deserves special attention. A significant correlation with geographical latitude was obtained only for the proportion of polymorphic loci at 99% criterion: r = 0.682, P < 0.05.
Cluster analysis, however, did not show distinct clusters of late-flowering populations and those with mixed time of flowering. This may be due to two reasons. First, mixed populations were represented both by spring and winter forms. Second, genetic characteristics of populations differing in the time of flowering were also affected by the climatic conditions in the northern marginal region of the species range.
The frequency of cross-pollination was estimated for each polymorphic locus. On average for all loci, this parameter was above zero in the populations, in which observed heterozygosity was recorded: in the most northern Kondopoga population (1.5%) and in three polymorphic for the time of flowering populations: Tsarevichi (1.1%), Pin’guba (2.4%), and Sheltozero
Table 4. Parameters of genetic diversity in Karelian populations of A. thaliana Geographical coordinates, N
Population Kondopoga Konchezero Kosalma Tsarevichi Suisar’ Pin’guba Chertov Stul Petrozavodsk Sheltozero Rybreka
62°12′ 62°08′ 62°01′ 62°01′ 61°58′ 61°53′ 61°50′ 61°48′ 61°22′ 61°16′ Mean
A
P95%
P99%
Hobs
1.8 (.3) 1.3 (.2) 1.9 (.4) 1.9 (.2) 1.9 (.3) 1.9 (.4) 1.3 (.2) 1.1 (.1) 1.3 (.2) 1.3 (.3) 1.6
62.5 12.5 62.5 50.0 50.0 50.0 25.0 0 25.0 12.5 35.0
62.5 25.0 62.5 75.0 62.5 62.5 37.5 12.5 25.0 12.5 43.7
0.011 (0.009) 0.000 (0.000) 0.000 (0.000) 0.004 (0.003) 0.000 (0.000) 0.010 (0.005) 0.000 (0.000) 0.000 (0.000) 0.002 (0.002) 0.000 (0.000) 0.003
H *exp 0.235 (0.071) 0.028 (0.022) 0.272 (0.082) 0.154 (0.062) 0.215 (0.084) 0.132 (0.060) 0.067 0.006 (0.06) 0.043 (0.029) 0.084 (0.084) 0.140 0.099 0.124
Correlation coefficient (r) with latitude
–
–
r = 0.68 P < 0.05
–
–
* Winter forms are shown at the left and mixed forms for the time of flowering are shown at the right. The Petrozavodsk population with a very low germinating capacity was not taken into account during calculations of the correlation coefficient. RUSSIAN JOURNAL OF GENETICS
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Table 5. Parameters of gene diversity (Nei’s statistics) in Karelian populations of A. thaliana Locus Got-1 Got-2 Idh-1 Acp-2 Acp-3 Acp-5 Est-1 Est-3 Mean
HT
HS
DST
GST
0.093 0.000 0.489 0.483 0.585 0.000 0.356 0.315 0.290
0.076 0.000 0.158 0.095 0.344 0.000 0.073 0.221 0.121
0.017 0.000 0.331 0.388 0.241 0.000 0.283 0.094 0.169
0.183 0.000 0.679 0.802 0.412 0.000 0.796 0.298 0.583
Note: Mean GST value was estimated from mean HT , HS , and DST values.
Based on allele frequencies, we estimated genetic similarity (IN) and genetic distances (DN) according to Nei [21]. The mean genetic similarity between all pairs of populations was 0.798, and mean genetic distances were 0.231. A correlation between genetic and geographical distances was not revealed. Genetic differentiation within and between populations was evaluated by Nei’s statistics of gene diversity [17]. The results are given in Table 5. The total gene diversity averaged over all loci HT was 0.290. The interpopulation component DST comprised the larger part of biodiversity. However, for three loci (Got-1, Acp-3, and Est-3), intrapopulation diversity (HS) was higher than the interpopulation component. In general, the relative value of interpopulation differentiation was 0.583. To compare our data with those obtained for other populations of A. thaliana [14] and for other self-pollinating species [22, 23], the mean values of the parameters obtained in this work and reported in the literature
are presented in Table 6. To facilitate comparison, A, HT, HS, and GST in Karelian populations (Table 6) were averaged only for polymorphic loci. The average number of alleles per polymorphic locus was similar to that in other populations of selfpollinating species. However, observed heterozygosity and cross-pollination frequency in the Karelian populations of A. thaliana were higher than in British populations and other self-pollinating species. Altered frequency of cross-pollination was observed in self-pollinating species under unfavorable environmental conditions: mountainous habitats, long day, and low temperatures in the Arctic [24]. In A. thaliana, this phenomenon was observed under elevated SO2 concentrations [25]. High frequency of cross-pollination revealed in Karelian A. thaliana populations may be characteristic of marginal populations. The polymorphism level (P99% = 43.7) was twice as high as the reported values given for comparison. High polymorphism may be associated, first, with a limited number of scored loci (eight), and second, with population-genetic peculiarities of the species in the northern part of its range. To be able to compare our data with those of Abbot and Gomes [14], we calculated the proportion of polymorphic loci in British populations of A. thaliana at eight isozyme loci (Idh-1, Got-1, Got-2, Acp-1, Acp-2, Acp-3, Est-1, and Est-2). The obtained values were similar to our estimates. As a result, the obtained mean value increased to 27.6% that was still lower than in Karelian populations. The comparison of gene differentiation in Karelian and British populations of A. thaliana showed that the distribution of genetic diversity within and between populations was similar and typical of self-pollinating species. However, the IN and DN mean values indicate that the interpopulation diversity in Karelian populations was higher than in other self-pollinating species (Table 6).
Table 6. Mean values of genetic diversity and cross-pollination frequency in populations of A. thaliana and other self-pollinating species Population A. thaliana, Karelian populations A. thaliana, British populations [14] Self-pollinating populations (14 populations) [22] Mean over 33 self-pollinating plant species [23]
Number of loci
I
H*T
H*S
G*ST
Cross-pollination frequency, %
0.798
0.387
0.161
0.527
0.56
16.5
0.00024 0.897
0.360
0.138
0.563
0.30
18.3
0.001
0.975
0.291
0.128
0.523
–
–
–
–
–
–
A
P99%
Hobs
8
1.57 2.26*
43.7
0.003
17 16.6
1.21 2.86* 2.26*
–
1.31
18.9
–
* Mean value estimated only for polymorphic loci. RUSSIAN JOURNAL OF GENETICS
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In conclusion, the population-genetic structure of A. thaliana growing in Karelia has several characteristic features. Despite the fact that the Karelian populations were mainly represented by late-flowering forms, several populations polymorphic for the time of flowering were found. Considerable population polymorphism at eight isozyme loci was revealed. The observed heterozygosity (Hobs = 0.003) and the proportion of polymorphic loci (P99% = 43.7%) in the Karelian populations of A. thaliana were more than twice as high as in populations from the central part of the species range and in populations of other self-pollinating species. The cross-pollination frequency was also higher in Karelian populations. A slight but significant correlation between the polymorphism level and latitude was shown (r = 0. 68; P < 0.05). In all probability, under the severe northern ecological conditions, the population polymorphism increases. In addition, the highest Hobs value was recorded in the most northern population. The distribution of genetic diversity within and between populations studied was typical of self-pollinating species, namely, its greater part was composed of the interpopulation component: GST = 0.583. ACKNOWLEDGMENTS The authors are grateful to P.V. Tikhov for fruitful discussion and help in statistical analysis of the data. REFERENCES 1. Kranz, A.R., Spontaneous and Induced Genetic Resources in Arabidopsis: Model and Tool to Study Certain Problems of Gene Exploration, AIS, 1976, no. 13, pp. 12–17. 2. Ratcliffe, D., Adaptation to Habitat in a Group of Annual Plants, J. Ecol., 1961, vol. 49, pp. 189–203. 3. Cetl, I., Genoclinal Character of Flowering-Time Variability in Arabidopsis thaliana (L.) Heynh., AIS, 1978, no. 5, pp. 92–109. 4. Mair, E., Zoologicheskii vid i evolyutsiya (Zoological Species and Evolution), Moscow: Mir, 1968. 5. Carson, H.L., Genetic Conditions Which Promote or Retard the Formation of Species, Cold Spring Harbor Symp. Quant. Biol., 1959, vol. 24, pp. 87–105. 6. Dobzhansky, T., Genetics and the Origin of Species, New York, 1951, 3rd ed. 7. Levontin, R.C., The Adaptations of Populations to Varying Environments, Cold Spring Harbor Symp. Quant. Biol., 1957, vol. 22, pp. 395–408. 8. Levontin, R.C., The Genetic Basis of Evolutionary Change, New York: Columbia Univ., 1974. 9. Korochkin, L.I., Serov, O.L., Pudovkin, A.I., et al., Genetika izofermentov (Genetics of Isozymes), Moscow: Nauka, 1977.
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10. Gomes, M.F. and Abbot, R.J., Genotypic Diversity in British Populations of Arabidopsis thaliana Based on a Survey of Isozyme Variation, AIS, 1987, no. 23, pp. 25– 30. 11. Fuglevitcz, A. and Kilian, A., Variability of Enzymatic Systems in Natural Populations of Arabidopsis thaliana in Poland, AIS, 1985, no. 22, pp. 87–90. 12. Grover, N.S., Characterization of Arabidopsis thaliana Ecotypes on the Basis of Genetic Variation at Ten Isozyme Loci, AIS, 1975, no. 12, pp. 19–21. 13. Jacobs, M. and Schwind, F., Biochemical Genetics of Acid Phosphatase Isozymes in Arabidopsis thaliana (L.) Heynh., AIS, 1976, no. 13, pp. 56–76. 14. Abbot, R.J. and Gomes, M.F., Population Genetic Structure and Outcrossing Rate of Arabidopsis thaliana (L.) Heynh., Heredity, 1989, vol. 62, no. 3, pp. 411–418. 15. Zhivotovsky, L.A., Statistical Methods for Analyzing Gene Frequencies in Natural Populations, Itogi Nauki Tekh., Ser.: Obshch. Genet., Moscow: VINITI, 1983, vol. 8, pp. 76–104. 16. Swofford, D.L. and Selander, R.B., BIOSYS-1: A FORTRAN Program for the Comprehensive Analysis of Electrophoretic Data in Population Genetics and Systematics, J. Hered., 1981, vol. 72, pp. 281–283. 17. Nei, M., Analysis of Gene Diversity in Subdivided Populations, Proc. Natl. Acad. Sci. USA, 1973, vol. 70, pp. 3321–3323. 18. Li, C.C., First Course in Population Genetics, Pacific Grove, California: Boxwood, 1976. 19. Kranz, A.R. and Kircheim, B., Genetic Resources in Arabidopsis, AIS, 1987, no. 24. 20. Cetl, I., Genetic Polymorphism for Alleles of FloweringTime in Natural Populations of Arabidopsis thaliana (L.) Heynh., AIS, 1990, no. 27, pp. 27–43. 21. Nei, M., Genetic Distance between Populations, Am. Nat., 1972, no. 106, p. 283. 22. Gottlieb, L.D., Electrophoretical Evidence and Plant Populations, Prog. Phytocem., 1981, vol. 7, pp. 1–46. 23. Hamrick, J.L., Linhart, Y.B., and Mitton, J.B., Relationship between Life History Characteristic and Electrophoretically Detectable Genetic Variation in Plants, Annu. Rev. Ecol. Syst., 1979, vol. 10, pp. 173–200. 24. Trofimovskaya, A.Ya., Yachmen’ (Barley), Leningrad: Kolos, 1972, p. 99. 25. Kilian, A. and Maluszynski, M., Genetic Variability of Arabidopsis thaliana Populations from Regions of Different Pollution Level, AIS, 1987, no. 25, pp. 57–66. 26. Weir, B.S., Allard, R.W., and Kahler, A.L., Further Analysis of Complex Allozymes Polymorphisms in Barley Population, Genetics, 1974, no. 3, pp. 911–919. 27. Zhivotovsky, L.A., Integratsiya poligennykh sistem v populyatsiyakh (Integration of Polygenic Systems in Populations), Moscow: Nauka, 1984.
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