<figure><img src="image_1.png" /></figure>

HAL
open science

# Electrophoretic variation in four french populations of domesticated rainbow trout (Salmo gairdneri)

René Guyomard

► To cite this version:

René Guyomard. Electrophoretic variation in four french populations of domesticated rainbow trout (Salmo gairdneri). Canadian journal of genetics and cytology. Journal canadien de genetique et de cytologie, 1981, 23, pp.33-47. 10.1139/g81-005. hal-01602926

HAL Id: hal-01602926

https://hal.science/hal-01602926

Submitted on 2 Jun 2020

HAL is a multi-disciplinary open access
archive for the deposit and dissemination of sci-
entific research documents, whether they are pub-
lished or not. The documents may come from
teaching and research institutions in France or
abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est
destinée au dépôt et à la diffusion de documents
scientifiques de niveau recherche, publiés ou non,
émanant des établissements d'enseignement et de
recherche français ou étrangers, des laboratoires
publics ou privés.

<figure><img src="image_2.png" /></figure>

Distributed under a Creative Commons Attribution - ShareAlike 4.0 International License
# ELECTROPHORETIC VARIATION IN FOUR FRENCH POPULATIONS OF DOMESTICATED RAINBOW TROUT (SALMO GAIRDNERI)

RENÉ GUYOMARD

Laboratoire de Physiologie des Poissons, Institut National de la Recherche Agronomique, 78350 Jouy en Josas, France

Electrophoretic variation of 12 enzyme systems, representing 26 loci in four strains of domesticated rainbow trout (*Salmo gairdneri* Richardson) were studied. The variances among and within strains, estimated from observed frequencies, were compared to data of other studies of biochemical polymorphism carried out in this species. These preliminary results show that a large proportion of the variability in rainbow trout is present in the French hatchery stocks studied and that isolation of these groups for several generations has not caused a significant genetic differentiation as compared to all the domesticated strains studied.

Nous avons examiné les variations électrophorétiques de 12 systèmes enzymatiques, représentant 26 locus chez 4 populations domestiques de Truite Arc-en-ciel (*Salmo gairdneri* Richardson). Les variabilités inter et intra-populations, estimées à partir des fréquences observées, ont été comparées aux données provenant d'autres études de polymorphisme biochimique réalisées sur la même espèce. De cette étude préliminaire, il ressort qu'une large part de la variabilité de l'espèce Truite Arc-en-ciel est présente dans les stocks piscicoles français étudiés et que l'isolement de ces groupes durant plusieurs générations ne s'est pas traduit par une différenciation génétique significative vis-à-vis de l'ensemble des populations domestiques étudiées.

## Introduction

The rainbow trout species (*Salmo gairdneri*) poses the following taxonomic problems at diverse levels.

- Its definition as compared to other "evolving lineages" of the genus *Salmo* in northwestern America. The polygenetic relations of the rainbow trout with such forms as *Salmo Aguabonita* and *S. apache* are still a controversial subject (Gold, 1977);

- Its genetic structure; at present, rainbow trout seems to be a heterogeneous species, including many geographic (*S. aquilarium*, *S. smaragdus*, *S. regalis*) and ecologic (anadromous, sedentary populations in rivers or lakes) forms. The existence of sympatric lake populations has been reported twice (Behnke, 1972).

All the breeding stocks which, as affected either by selection due to domestication or by random sampling (initial or at each breeding cycle), have differentiated into one or more original sub-units should be added to these natural populations.

Among the various techniques available for phylogenetic studies, those using the genetic variations in proteins have proven particularly useful; they have been widely employed in studies in the genus *Salmo* in northwestern America, indicating the phylogenetic relations within that group (Gold, 1977; Allendorf and Utter, 1974).

Work carried out on the migratory form of rainbow trout (Allendorf, 1975; Utter and Allendorf, 1977) has revealed two genetically differentiation groups corresponding to two distinct geographical areas, one coastal and the other inland. Those authors invalidated the conventional divisions according to seasonal return to the river and reported no genetic diversity between anadromous and resident forms in the same river (Utter and Allendorf, 1977).
The rainbow trout was introduced into Europe some 100 years ago (Mac
Crimmon, 1971); we wished to determine what part of the genetic variability of
the species was thus introduced and maintained. This paper reports the electrophoretic
variants of different enzyme systems in some rainbow trout populations used in
French fish culture and attempts to answer three questions:

1) what is the proportion of heterozygosity in these populations

2) what is their degree of differentiation

3) what is their biochemical polymorphism as compared to the main body of the species.

In this article, we present some preliminary results obtained in four of these
populations.

**Material and Methods**

Strains

The four strains studied were taken from three different fish farms and were chosen
from a list of farms whose breeding stocks had not been renewed from the outside for at
least ten generations. No more data are available about the history of these strains. In one
farm, we sampled two different groups, one in 1977 and the other in 1978. The one to two-
year old fish were killed and stored at -20°C, and their tissues were tested during the next
two weeks. These four strains are named S<sub>1</sub>-77, S<sub>1</sub>-78, S<sub>2</sub>, S<sub>3</sub> and the sizes of samples from
these ones are respectively 63, 40, 60 and 40.

Extraction

The extracts were obtained after grinding in a 0.01M phosphate buffer pH 7.4 (weight/volume). The ground heart, eye and muscle were centrifuged at 4000 xg for 30 min, and sometimes re-centrifuged; liver extracts were centrifuged at 26000 xg for 1.5 h. All manipulations were carried out at 4°C.

Electrophoresis

300 ml of 12% Connaught starch gel were poured into 6 × 170 × 225 mm molds. The clear supernatants were individually drawn into 6 × 6 mm squares of Whatman 3<sup>M</sup> paper inserted into the gel. Buffers, electrophoretic conditions and the staining solutions of each system are shown in Table I. The gels were refrigerated during migration with ice packs.

Designation of Alleles

The locus designation adopted here conforms to the conventional nomenclature (Allendorf, 1975; Busack *et al.*, 1979). The most frequent alleles were arbitrarily designated by mobility 1.00.

Measurement of Genetic Distance and Proportion of Polymorphism

We used the formula proposed by Nei (1975). If xij is the frequency of allele i for a
locus in a population j, gene diversity is expressed by Hj = 1 - Σ xij which is equivalent
to average heterozygosity, assuming Hardy-Weinberg genotype proportions. Genetic distance
between populations j and j' is measured by Djj' = 1/2 Σ (xij - xij')². The overall genic diver-
sity of a group of populations is Ht = 1 - Σ xij² with x_i = Σ w_j x_{ij} where w_j is the coeffici-
ent of the weight of the population j. In this paper we used the same coefficient for each
population (w_j = 1/N) with N representing the number of populations. Extensions to several
loci are immediate.
<table><caption>TABLE I
Electrophoretic and staining techniques</caption><thead><tr><th>Systems</th><th>Examined tissue</th><th>Electrophoretic conditions</th><th>Revelation</th></tr></thead><tbody><tr><td>Alcohol dehydrogenase (ADH)</td><td>Liver</td><td>Ridgway buffer (Ridgway et al., 1970); 60 Ma, voltage max 350 volts; migration during 3 h</td><td>Ethanol 5 ml; NAD 15 mg; NBT 5 mg; PMS 5 mg; 0.05M Tris-HCl, pH 8.00 : 100 ml</td></tr><tr><td>Creatine kinase (CK)</td><td>Muscle</td><td>Ridgway buffer</td><td>1) Specific staining : creatine kinase, disodium salt 20 mg; NBT 5 mg; PMS 10 mg; NADP 5 mg; adenosine 5' diphosphate 30 mg; glucose 20 mg; hexokinase 100 units; g-6-pdh 20 units; 0.05M Tris-HCl, pH 8.00 : 100 ml 2) Amidoblack staining</td></tr><tr><td>Glutamate oxaloacetate transaminase (GOT)</td><td>Heart</td><td>Morpholino-citrate buffer (Clayton and Tretiak, 1972) 65mA; 320 volts during 5 h</td><td>Aspartic acid : 100 mg; Pyridoxal 5' phosphate 10 mg; Fast blue BB salt 100 mg; α-keto-glutaric acid 80 mg; 0.2M Tris-HCl pH 8.00 : 100 ml</td></tr><tr><td>α-glycero-phosphate dehydrogenase (GPDH)</td><td>Muscle</td><td>0.1M Tris-phosphate pH 7.4 90 mA; 150 volts during 5 h</td><td>DLα glycerophosphate, disodium salt 80 mg; PMS 5 mg; NAD 25 mg; NBT 5 mg; EDTA Trisodium salt 16 mg; 0.05M Tris-HCl, pH 8.00 : 100 ml</td></tr><tr><td>Isocitrate dehydrogenase (IDH-s)</td><td>Liver</td><td>1) 0.1M Tris-phosphate pH 7.4 90 mA; 150 volts during 5 h<br>2) sodium citrate buffer (Cross and Payne, 1977); 100 mA; 130 volts during 9 h</td><td>D-L-isocitric acid, trisodium salt 30 mg; NADP 5 mg; NBT 5 mg; PMS 5 mg; MgSO4 40 mg; 0.05M Tris-HCl pH 8.00 : 100 ml</td></tr><tr><td>Mitochondrial isocitrate dehydrogenase (IDH-m)</td><td>Heart</td><td>0.1M Tris-phosphate pH 7.4 90 mA, 150 volts during 5 h</td><td>D-L-isocitric acid, trisodium salt 30 mg; NADP 5 mg; NBT 5 mg; PMS 5 mg; MgSO4 40 mg; 0.05M Tris-HCl pH 8.00 : 100 ml</td></tr><tr><td>Lactate dehydrogenase (LDH)</td><td>Muscle</td><td>Ridgway buffer</td><td>Lactic acid, lithium salt 200 mg; NAD 15 mg; NBT 5 mg; PMS 5 mg; Tris-HCl pH 8.00 : 100 ml</td></tr><tr><td>Malate dehydrogenase (MDH)</td><td>Liver<br>Muscle<br>Heart</td><td>Morpholino-citrate buffer (Clayton and Tretiak, 1972) 65 mA; 320 volts during 5 h</td><td>L-Malic acid, monosodium salt 500 mg; NAD 40 mg; NBT 20 mg; PMS 5 mg; 0.15M Triethano-lamine-HCl pH 9.0 : 100 ml</td></tr><tr><td>Phosphoglucose isomerase (PGI)</td><td>Muscle</td><td>0.1M Tris-phosphate pH 7.4 90 mA; 150 volts during 5 h</td><td>Fructose-6-phosphate 12.5 mg; NADP 0.8 mg; NBT 1.5 mg; PMS 1.5 mg; G-6-PDH 25 units; 0.05M Tris-HCl pH 8.00 : 4 ml. 10 × 22.5 cm rectangle of Whatman n° 1 paper, impregnated with this solution, is applied on the gel</td></tr><tr><td>Phosphoglucomutase (PGM)</td><td>Muscle</td><td>0.1M Tris-phosphate pH 7.4 90 mA, 150 volts during 5 h</td><td>Same staining procedure; 15 mg of Glucose-1-phosphate instead of F-6-phosphate</td></tr><tr><td>Transferrin (Tf)</td><td>Serum</td><td>Ridgway buffer</td><td>Amidoblack</td></tr><tr><td>Superoxide dismutase (S.O.D.)</td><td>Liver</td><td>Ridgway buffer</td><td>Stained in the same time as ADH</td></tr></tbody></table>

Estimation of Allelic Frequencies

An extensive gene duplication was found in salmonid species, confirming that they have been subjected to a tetraploidization during their evolution. These species now have a disomic
mode of inheritance; at the molecular level this means that in some tissues, some enzyme systems are coded by two loci. Since the allelic relations between these four genes and their assignment to each locus could not always be determined on the electrophoregrams, sometimes the genotypes could be only partially ascertained for two loci of a duplicate. We had this problem with the glutamic oxaloacetic transaminase (GOT), malate dehydrogenase (MDH) and isocitrate dehydrogenase (IDH) systems, and therefore adopted the following rule: if $n_{ij}$ was the total observed number of genes of a given allelic type i and $n_j$ the total number of genes sampled, the genetic distance, computed with these two loci was minimal if the frequency $\frac{n_{ij}}{2n_j}$ was assigned to the allele i at each of the two loci. This was the case of

<figure><img src="image_3.png" /><figcaption>FIG. 1. 1a: GOT in heart; 1b: details of polymorphism corresponding to loci Got1 and 2. FIG. 2. MDH in heart. FIG. 3. IDH-m in heart. FIG. 4. Idh-s 3 and 4; presumed genotypes: 1, 8, 10, 17, 19, 20. Idh 3-4 (1.00/1.00/1.00/0.38); 2: Idh 3-4 (1.00/1.00/1.00/0.62); 3, 7, 9, 14: Idh 3-4 (1.00/1.00/0.38/0.38); 4, 12, 13: Idh 3-4 (1.00/1.00/0.38/1.31); 5: Idh 3-4 (1.00/1.31/0.69/0.69); 6: Idh 3-4 (1.00/0.69/0.69/0.38); 11, 15: Idh 3-4 (1.00/1.00/1.00/1.00); 16: Idh 3-4 (1.00/1.00/1.00/1.31); 18: Idh 3-4 (1.00/1.00/0.69/0.69).</figcaption></figure>
<figure><img src="image_4.png" /></figure>

<figure><img src="image_5.png" /><figcaption>FIG. 5. Genetic interpretation of FIG. 1a. 1, 2 and 5: Got 1-2 (1.00/1.00/1.00/1.17); 3: Got 1-2 (1.00/1.00/1.00/0.70); 4: Got 1-2 (1.00/1.00/1.17/1.17).</figcaption></figure>

### IDH and MDH.
The whole of the values $\frac{n_{ij}}{2n_j}$ ($j = 1, ..., n$) minimized the mean genetic distance $\frac{1}{N^2} \sum_{j=1}^{n} \sum_{w_j} w_j D_{jj'}$, where N is the number of populations, but maximized $\sum_j H_j w_j$ as well as the overall genetic diversity $H_T$. The degree of heterogeneity of the populations group $\frac{\sum_{j=1}^{n} \sum_{w_j} w_j D_{jj'}}{N^2 H_T}$ was also minimum, but the dendrogram of those values did not necessarily correspond to that obtained from the true frequencies. For GOT, we considered the possibility that only one of the two loci might be polymorphic; both hypotheses (one or two polymorphic loci) were tested using the law of Hardy-Weinberg.

### Comparison with Results in the Literature

Two papers (Allendorf, 1975; Busack *et al.*, 1979) have furnished data on nine stocks from American fish farms; we have compared these with our results. For the between-population comparisons, we used only the loci common to the three studies. For each one, we accepted the homologies of the alleles most frequently described in the three articles; this was well corroborated by the electrophoretic mobilities of other alleles. For both the MDH and IDH systems, we regrouped some observed alleles which could not be differentiated in the other two studies (see Results).

### Construction of Dendrograms

The dendrogram indicating the genetic distances between the 13 populations was constructed using a Fortran program provided by Dr. Neï.
<figure><img src="image_6.png" /><figcaption>FIG. 6. Genetic interpretation of Fig. 3. 1 : Idh-2 (1.00/1.00); 2 : Idh-2 (1.00/1.20).</figcaption></figure>

## Results

### Description and Interpretation of Electrophoregrams

We studied 11 enzymatic systems, and the electrophoregrams obtained agreed with the results of the other two authors (Allendorf, 1975; Busack *et al.*, 1979). Table II shows the models currently used in the genetic determinism of those systems and preferentially indicated the references mentioning "genetic tests" carried out on rainbow trout species. The other methods sometimes used are genetic studies in phylogenetically close species and the distribution of enzymatic forms in the various tissues. Figures 1, 2, 3, 4, 5, 6 and 7 illustrate some of the variants found.

The electrophoretic behavior of GOT deserves attention; in Ridgway and Trisphosphate buffers, the system was monomorphic. This agrees with the observations of Allendorf (1975) and Busack *et al.* (1979). However, at pH 6.1 extreme variations resembling those of MDH-B and suggesting a genetic control of two loci, were found (Figs. 1a and 1b). Table II also presents results on the duplicated or unduplicated state of the different loci studied. Whether or not duplication is found partially depends on technique sensitivity, as illustrated for α-glycerophosphate dehydrogenase (α-GPDH) (Allendorf, 1975). We estimated a total of 26 loci for the 11 enzymatic systems.

### Allelic Frequencies

Table III shows the alleles found and their computed frequencies at each locus. For IDH-s and MDH-B, each allele was presumed to have the same frequency at the two loci coding for each system (minimal genetic distance). For GOT according to the test of the two hypotheses (minimization of genetic distance and only one polymorphic locus) using the law of Hardy-Weinberg, we retained the second one in agreement with this law (Table IV).

The distribution of the allelic frequencies in each stock, followed a conventional pattern: at most loci a high frequency allele common to all the populations was found.

We usually observed a larger number of alleles than mentioned in the other two studies of rainbow trout populations (Allendorf, 1975; Busack *et al.*, 1979).
<table><caption>TABLE II
Genetic determinism of studied enzymatic systems</caption><thead><tr><th>Systems</th><th>Enzyme structure</th><th>Genetic determinism</th><th>Duplication</th><th>References</th></tr></thead><tbody><tr><td>APH</td><td>dimeric</td><td>1 locus</td><td>no</td><td>Allendorf (1975): mendelian inheritance studies on rainbow trout</td></tr><tr><td>CK</td><td>monomeric</td><td>2 loci:<br>Cpk-1 variable<br>Cpk-2 fixed</td><td>yes, disomic<br>heredity</td><td>Allendorf (1975): mendelian inheritance studies on rainbow trout</td></tr><tr><td>GOT</td><td>dimeric</td><td>2 loci:<br>Got-1 fixed<br>Got-2 variable</td><td>idem</td><td>Allendorf (1975): mendelian inheritance studies on rainbow trout</td></tr><tr><td>α-GPDH</td><td>idem</td><td>2 loci:<br>αgpdh-1 fixed<br>αgpdh-2 variable</td><td>idem</td><td>Utter et al. (1973); May et al. (1979); mendelian inheritance tests on brook<br>trout (Salvelinus fontinalis) and lake trout (Salvelinus namaycush) × brook<br>trout</td></tr><tr><td>IDH-s</td><td>idem</td><td>2 loci: Idh-3 and<br>Idh-4 variable</td><td>idem</td><td>Allendorf and Utter (1973): inheritance studies on rainbow trout</td></tr><tr><td>IDH-m</td><td>idem</td><td>2 loci: Idh-1<br>fixed, Idh-1<br>variable</td><td>idem</td><td></td></tr><tr><td>LDH-M</td><td>tetrameric</td><td>2 loci: Ldh-1 and<br>Ldh-2 fixed</td><td>idem</td><td>Bailey et al. (1976)</td></tr><tr><td>LDH-H</td><td>idem</td><td>2 loci: Ldh-3 and<br>Ldh-4 variable</td><td>idem</td><td>Wright et al. (1975): inheritance studies on rainbow trout</td></tr><tr><td>LDH-E</td><td>idem</td><td>1 locus, Ldh-5<br>variable</td><td>no</td><td>Wright et al. (1975): inheritance studies on rainbow trout</td></tr><tr><td>MDH-A</td><td>dimeric</td><td>2 loci: Mdh 1 and 2</td><td>yes</td><td>Allendorf (1975): inheritance studies on rainbow trout</td></tr><tr><td>MDH-B</td><td>idem</td><td>2 loci: Mdh 3 and<br>4, variable</td><td>yes disomic<br>heredity</td><td>Bailey et al. (1970): inheritance studies on rainbow trout</td></tr><tr><td>PGI</td><td>idem</td><td>3 loci: Pgi 1,<br>2 and 3</td><td>idem for Pgi-1<br>and 2, Pgi-3<br>not duplicated</td><td>Avise and Kitto (1973); Schmidtke et al. (1975); May et al. (1979):<br>inheritance studies on brook trout</td></tr><tr><td>PGM</td><td>monomeric</td><td>1 locus variable</td><td>no</td><td>Roberts et al. (1979); Utter et al. (1973): inheritance studies on sockeye<br>salmon (Oncorhynchus nerka)</td></tr><tr><td>TO</td><td>dimeric</td><td>idem</td><td>idem</td><td>Utter et al. (1973): inheritance studies on rainbow trout</td></tr><tr><td>Tf</td><td>monomeric</td><td>idem</td><td>idem</td><td>Utter et al. (1973): inheritance studies on rainbow trout</td></tr></tbody></table>
<figure><img src="image_7.png" /><figcaption>FIG. 7. Genetic interpretation of Fig. 5 for anodal variations. Cathodal part should correspond to mitochondrial MDH.</figcaption></figure>

1 : Mdh 3-4 (1.00/1.00/1.00/1.00); 2 : Mdh 3-4 (1.00/1.00/1.00/0.96);

3 : Mdh 3-4 (1.00/1.00/1.00/0.79); 4 : Pdh 3-4 (1.00/1.00/1.00/0.70)

This is certainly not related to the sample size of the populations, but to true
differences between the frequencies or more likely to the electrophoretic techniques
used.

The eight loci, A-gpdh 1 and 2, Idh 3 and 4, Mdh 3 and 4, Pgm and Sod,
common to the three studies, were used for subsequent comparisons. These eight
loci give highly biaised absolute values, but we are mainly interested in relative
comparisons. Four relatively variable systems (IDH-m, GOT, CK and p-ALB), either
studied under very different electrophoretic conditions or not examined in the overall
populations, were discarded.

For IDH-s and MDH-B, we regrouped the following alleles; Idh (1.31-1) and
(1.31-2), Idh (0.69) and (0.62), Mdh (1.00) and (0.96), Mdh (0.79) and Mdh (0.70).
The allelic frequencies for IDH-s reported by Allendorf (1975) had to be revised,
that author having hypothesized a single variable locus for the system (Appendix
Table I). Finally, we tested the panmictic deviation at some loci; the χ² values
obtained are shown in Table IV. The gene diversity (Table V) and genetic distances
(Table VI) were computed according to Neï (1975). The distance dendrogram is
shown in Fig. 8.
<table><caption>TABLE III
Allelic frequencies</caption><thead><tr><th>System locus<br>and alleles</th><th>strain 1-77 n=63</th><th>strain 1-78 n=40</th><th>strain <sup>2</sup> n=60</th><th>strain <sup>3</sup> n=40</th></tr></thead><tbody><tr><td>ADH Adh</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>CPK Cpk1 (1.00)</td><td>-</td><td>0.975</td><td>-</td><td>0.885</td></tr><tr><td>Cpk1 (0.48)</td><td>-</td><td>0.025</td><td>-</td><td>0.115</td></tr><tr><td>Cpk2</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>GOT Got-1</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>Got-2(1.00)</td><td>-</td><td>0.562 (X<sup>2</sup><sub>1</sub> = 0.74)</td><td>-</td><td>0.650 (X<sup>2</sup><sub>1</sub> = 0.58)</td></tr><tr><td>Got-2(1.17)</td><td>-</td><td>0.438</td><td>-</td><td>0.350</td></tr><tr><td>αGPDH αgpdh1</td><td>fixed</td><td>fixed</td><td>fixed</td><td>fixed</td></tr><tr><td>αgpdh2(a)</td><td>0.984</td><td>1.000</td><td>1.000</td><td>0.975</td></tr><tr><td>αpdh2(b)</td><td>0.016</td><td>0.000</td><td>0.000</td><td>0.025</td></tr><tr><td>IDH-s Idh 3 and 4</td><td></td><td></td><td></td><td></td></tr><tr><td>Idh (1.00)</td><td>0.595</td><td>0.587</td><td>0.646</td><td>0.612</td></tr><tr><td>Idh (0.38)</td><td>0.230</td><td>0.250</td><td>0.154</td><td>0.244</td></tr><tr><td>Idh (0.62)</td><td>0.024</td><td>0.013</td><td>0.071</td><td>0.013</td></tr><tr><td>(1) Idh (1.31-1)</td><td>0.091</td><td>0.081</td><td>0.046</td><td>0.031</td></tr><tr><td>(1) Idh (1.31-2)</td><td>0.012</td><td>0.006</td><td>0.054</td><td>0.062</td></tr><tr><td>Idh (0.69)</td><td>0.048</td><td>0.063</td><td>0.029</td><td>0.038</td></tr><tr><td>IDH-m Idh1</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>Idh2(1.00)</td><td>-</td><td>0.850</td><td>-</td><td>0.862</td></tr><tr><td>Idh2(1.20)</td><td>-</td><td>0.150</td><td>-</td><td>0.138</td></tr><tr><td>LDH-M Ldh 1 and 2</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>LDH-H Ldh 3 and 4</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>LDH-E Ldh 5</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>MDH-A Mdh 1</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>MDH-B Mdh 3 and 4:</td><td></td><td></td><td></td><td></td></tr><tr><td>Mdh (1.00)</td><td>0.716</td><td>0.719</td><td>0.916</td><td>0.731</td></tr><tr><td>Mdh (0.79)</td><td>0.156</td><td>0.156</td><td>0.071</td><td>0.160</td></tr><tr><td>Mdh (0.70)</td><td>0.083</td><td>0.081</td><td>0.013</td><td>0.045</td></tr><tr><td>Mdh (0.96)</td><td>0.045</td><td>0.044</td><td>0.000</td><td>0.064</td></tr><tr><td>PGI Pgi 1,2 and 3</td><td>-</td><td>fixed</td><td>-</td><td>fixed</td></tr><tr><td>PGM Pgm (1.00)</td><td>0.700 (X<sup>2</sup><sub>1</sub>=0.57)</td><td>0.737 (X<sup>2</sup><sub>1</sub>=2.06)</td><td>0.983</td><td>0.810</td></tr><tr><td>Pgm (0.70)</td><td>0.300</td><td>0.263</td><td>0.017</td><td>0.190</td></tr><tr><td>TO To (1.00)</td><td>0.340 (X<sup>2</sup><sub>1</sub>=0.12)</td><td>0.275 (X<sup>2</sup><sub>1</sub>=1.01)</td><td>0.640 (X<sup>2</sup><sub>1</sub>=9.1*)</td><td>0.350 (X<sup>2</sup><sub>1</sub>=0.01)</td></tr><tr><td>To (1.61)</td><td>0.660</td><td>0.712</td><td>0.280</td><td>0.637</td></tr><tr><td>To (0.39)</td><td>0.000</td><td>0.013</td><td>0.080</td><td>0.013</td></tr><tr><td>TFN Tfn (a)</td><td>0.940</td><td>0.962</td><td>0.992</td><td>0.975</td></tr><tr><td>Tfn (b)</td><td>0.060</td><td>0.038</td><td>0.008</td><td>0.025</td></tr></tbody></table>

Distinction between Idh (1.31-1) and Idh (1.31-2) is based on utilisation of two buffers : 0.1M Tris-phosphate pH 7.4 and 0.04M Sodium Citrate pH 5.

$X_1^2$ values correspond to tests of Hardy-Weinberg law * 0.001<p<0.005

n = sample size

## Discussion

### Intrapopulation Variability

The four stocks studied ($S_1$-77, $S_1$-78, $S_2$, $S_3$) showed a higher mean proportion of polymorphism than other populations. On account of the precautions taken, it would seem that the effect of the electrophoretic techniques can be eliminated, but no more precise reasons can be given for this variability of heterozygosity.

A highly polymorphic domesticated population may result from a mixture of natural populations which are less polymorphic but genetically differentiated or a high polymorphic natural population; this possibility cannot be rejected. Some steel-head populations are very polymorphic (computed from the data of Allendorf, 1975
<caption>TABLE IV
Test of Hardy-Weinberg law for GOT under two hypotheses</caption>

1) one locus is fixed 2) allele b is present at the two loci with the same frequency

<table><thead><tr><th rowspan="2"></th><th colspan="2">Observed</th><th colspan="2">Expected (hypothesis 1)</th><th colspan="2">Expected (hypothesis 2)</th></tr><tr><th>Strain 1-78</th><th>Strain 3</th><th>Strain 1-78</th><th>Strain 3</th><th>Strain 1-78</th><th>Strain 3</th></tr></thead><tbody><tr><td>aaaa</td><td>14</td><td>18</td><td>12.5</td><td>16.9</td><td>3.93</td><td>7.14</td></tr><tr><td>aaab</td><td>17</td><td>16</td><td>19.7</td><td>18.2</td><td>12.36</td><td>15.38</td></tr><tr><td>aabb</td><td>9</td><td>6</td><td>7.8</td><td>4.9</td><td>14.57</td><td>12.42</td></tr><tr><td>abbb</td><td>0</td><td>0</td><td>0</td><td>0</td><td>7.63</td><td>4.46</td></tr><tr><td>bbbb</td><td>0</td><td>0</td><td>0</td><td>0</td><td>1.50</td><td>0.6</td></tr></tbody><tfoot><tr><td colspan="2">X<sup>2</sup> =</td><td></td><td>X<sup>2</sup><sub>1</sub>=0.74 X<sup>2</sup><sub>2</sub>=0.52</td><td>X<sup>2</sup><sub>3</sub>=38.8*X<sup>2</sup><sub>4</sub>=24.92*</td><td></td><td></td></tr></tfoot></table>

for the same subsample of eight loci, H = 0.31 in a natural steelhead population, and the resident forms which do not seem to be genetically differentiated from the migratory ones might exhibit the same degree of polymorphism.

Although it is impossible to choose between these two possibilities, we can conclude that introduction and breeding have not led to a reduction of the intra-population genetic variability of these stocks. Such processes, however, seem to operate in some populations, as in the stock of the University of Washington (Allendorf, 1975).

Table V shows the proportion of heterozygosity of stocks S₁-78 and S₃ computed from the whole of the systems studied. The values in parentheses correspond to monomorphic Mdh₄ and Idh₄ loci, all the variations of the MDH-A and IDH-s systems being reported to the Mdh₃ and Idh₃ loci, respectively.

Allendorf and Utter (1978) have demonstrated that rainbow trout are highly polymorphic as compared to other salmonids. Comparisons with other species entail some corrections; the mean value of intrapopulation variability (0.22) was computed for eight loci, seven of which were very polymorphic in the rainbow trout. Using

<table><caption>TABLE V
Heterozygosity levels</caption><thead><tr><th></th><th>8 loci</th><th>25 loci</th></tr></thead><tbody><tr><td>S<sub>1</sub>-77</td><td>0.348</td><td></td></tr><tr><td>S<sub>1</sub>-78</td><td>0.336</td><td>0.149 (0.125)</td></tr><tr><td>S<sub>2</sub></td><td>0.241</td><td></td></tr><tr><td>S<sub>3</sub></td><td>0.324</td><td>0.149 (0.134)</td></tr><tr><td>Univ. of Washington*</td><td>0.086</td><td></td></tr><tr><td>Chamber Creek*</td><td>0.228</td><td></td></tr><tr><td>Mac Creary*</td><td>0.210</td><td></td></tr><tr><td>West Virginia*</td><td>0.088</td><td></td></tr><tr><td>RTH**</td><td>0.171</td><td></td></tr><tr><td>RTHX**</td><td>0.174</td><td></td></tr><tr><td>RTS**</td><td>0.214</td><td></td></tr><tr><td>RTW**</td><td>0.295</td><td></td></tr><tr><td>RTV**</td><td>0.150</td><td></td></tr></tbody></table>

<footnote>*: <0.0005</footnote>

<footnote>*Allendorf (1975).</footnote>

<footnote>**Busack *et al.* (1979).</footnote>
<figure><img src="image_8.png" /><figcaption>FIG. 8. Dendrogram of genetic distances between 13 populations of domesticated rainbow trout.</figcaption></figure>

some of Allendorf's data (1975) taken from a more representative sample of the whole of the loci generally used for estimating the proportion of heterozygosity, we obtained a correct estimate of $H_c = 0.063$. Other vertebrates (28 species in Fuerst *et al.*, 1977) shown a mean value of 0.038 (SD = 0.033) after elimination of very variable esterases ($H_{esterases} = 0.134$ for the 28 species) which Allendorf did not study.

## Interpopulation Variability

Samples S₁-78 and S₁-77, representing two successive groups from the same fish farms, were almost identical. This frequency stability, which should be followed through several generations, has been reported several times (Altukhov *et al.*, 1975; Busack *et al.*, 1979), and shows the repeatability of the method, the representative nature of the samples and the advantage of this technique as a tool for the genetic description of species and populations.

Table VII gives the analysis of overall genetic variance into mean intrapopulation genetic diversity and mean interpopulation genetic distance for the whole of the
<table><caption>TABLE VI
Genetic distances matrix</caption><thead><tr><th></th><th>S<sub>1</sub>-77</th><th>S<sub>1</sub>-78</th><th>S<sub>2</sub></th><th>S<sub>3</sub></th><th>Univ. of Washington</th><th>Chamber Creek</th><th>Mac Creary</th><th>West Virginia</th><th>RTH</th><th>RTHX</th><th>RTS</th><th>RTW</th></tr></thead><tbody><tr><td>S<sub>1</sub>-78</td><td>0.00074</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>S<sub>2</sub></td><td>0.03222</td><td>0.03537</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>S<sub>3</sub></td><td>0.00199</td><td>0.00187</td><td>0.02244</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Univ. of Washington</td><td>0.08566</td><td>0.09356</td><td>0.02039</td><td>0.07180</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Chamber Creek</td><td>0.01723</td><td>0.01984</td><td>0.02809</td><td>0.01586</td><td>0.05575</td><td></td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>Mac Creary</td><td>0.03041</td><td>0.03422</td><td>0.01958</td><td>0.02421</td><td>0.04200</td><td>0.01395</td><td></td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>West Virginia</td><td>0.07047</td><td>0.07624</td><td>0.01492</td><td>0.05664</td><td>0.00402</td><td>0.03935</td><td>0.02380</td><td></td><td></td><td></td><td></td><td></td></tr><tr><td>RTH</td><td>0.03392</td><td>0.02976</td><td>0.07363</td><td>0.03393</td><td>0.14328</td><td>0.03085</td><td>0.04641</td><td>0.10914</td><td></td><td></td><td></td><td></td></tr><tr><td>RTHX</td><td>0.03601</td><td>0.03345</td><td>0.08196</td><td>0.03914</td><td>0.14881</td><td>0.02855</td><td>0.04777</td><td>0.11464</td><td>0.00216</td><td></td><td></td><td></td></tr><tr><td>RTS</td><td>0.03364</td><td>0.03793</td><td>0.00318</td><td>0.02518</td><td>0.01507</td><td>0.02115</td><td>0.01382</td><td>0.00857</td><td>0.07179</td><td>0.07709</td><td></td><td></td></tr><tr><td>RTW</td><td>0.06608</td><td>0.07270</td><td>0.03431</td><td>0.05959</td><td>0.06079</td><td>0.09349</td><td>0.08266</td><td>0.07069</td><td>0.06102</td><td>0.17174</td><td>0.04840</td><td></td></tr><tr><td>RTV</td><td>0.05003</td><td>0.05183</td><td>0.03038</td><td>0.03834</td><td>0.04991</td><td>0.02744</td><td>0.00831</td><td>0.02875</td><td>0.05297</td><td>0.05878</td><td>0.02446</td><td>0.11060</td></tr></tbody></table>
<caption>TABLE VII</caption>

$$ \text{Analysis of genic diversity } H = \frac{\sum_j H_j}{N} + \frac{1}{N^2} \sum_{jj'} \sum D_{jj'} $$

<table><thead><tr><th>Population grouping</th><th>Genic diversity<br>intrapopulation <sup>ΣHj</sup>/<sub>N</sub></th><th>Average genetic<br>distance <sup>ΣΣDjj'</sup>/<sub>N<sup>2</sup></sub></th></tr></thead><tbody><tr><td>H<sub>T</sub> : (all 13 populations)</td><td>0.210</td><td>0.048</td></tr><tr><td>H<sub>1</sub> : (the 4 French populations)</td><td>0.302</td><td>0.013</td></tr><tr><td>H<sub>2</sub> : (the other 9 populations)</td><td>0.179</td><td>0.052</td></tr><tr><td>H'<sub>T</sub> : (French populations and<br>American populations<br>considered as two groups)</td><td>0.270</td><td>0.006</td></tr></tbody></table>

13 populations (H<sub>T</sub>), the four stocks studied (H<sub>1</sub>), the other nine populations (H<sub>2</sub>), as well as the classification obtained when the French populations and the American populations were considered as two different units (H'<sub>T</sub>).

The mean genetic distance within the groups S<sub>1</sub>, S<sub>2</sub> and S<sub>3</sub> is lower (0.013 vs. 0.052), indicating strong homogeneity. However, examination of H'<sub>T</sub> does not reveal any significant differentiation between the two groups. This is illustrated by the genetic distribution dendrogram in Fig. 8.

The constitution of French stocks began about 100 years ago with populations introduced from North America. These introductions, which are still in progress either directly or by the intermediary of some European country or other continents, are difficult to trace. However, it would appear from this preliminary study that a large part of the genetic variability of the rainbow trout species is present in the stocks studied and that the isolation of these groups for several generations has not caused significant genetic differentiation, as compared to all the domesticated strains studied.

## References

* Allendorf, F. W. 1975. Genetic variability in a species possessing extensive gene duplication; Genetic interpretation of duplicate loci and examination of genetic variation in populations of rainbow trout. Ph.D. Thesis, Univ. Washington, Seattle. pp. 98.

* Allendorf, F. W. and Utter, F. M. 1973. Gene duplication within the family salmonidae; Disomic inheritance of two loci reported to be tetrasomic in rainbow trout. Genetics, 74: 647-654.

* Allendorf, F. W. and Utter, F. M. 1974. Biochemical genetic systematics of the genus salmon. Anim. Blood Groups Biochem. Genet. 5(Suppl. 1): 33. Abstr.

* Allendorf, F. W. and Utter, F. M. 1978. Population genetics of fish. In Fish physiology. Edited by W. S. Hoar, D. J. Randall and J. R. Breet. Academic Press, New York. 8: 407-545.

* Altukhov, Y. P., Salmenkova, F. A., Konovalov, S. M. and Padoukin, A. I. 1975. Stationary distributions of frequencies of lactate dehydrogenase and phosphoglucomutase genes in population system of local fish stock *Oncorhynchus nerka*. I. Stability of the stock in generations under simultaneous variability of subpopulations making up their structure. Genetika, 11: 44-53.

* Avise, J. C. and Kitto, G. B. 1973. Phosphoglucose isomerase gene duplication in the bony fishes: An evolutionary history. Biochem. Genet. 8: 113-131.

* Bailey, G. S., Wilson, A. G., Halver, J. E. and Johnson, C. L. 1970. Multiple forms of supernatant malate dehydrogenase in salmonid fishes: Biochemical, immunological and genetic studies. J. Biol. Chem. 245: 5927-5940.

* Bailey, G. S., Tsuyuki, H. and Wilson, A. C. 1976. The number of genes for lactate dehydrogenase in salmonid fishes. J. Fish. Res. Board Can. 33: 760-767.

* Behnke, R. J. 1972. The systematics of salmonid fishes or recently glaciated lakes. I. Subfamily salmonidae. J. Fish. Res. Board Can. 29: 639-671.
Busack, C. A., Halliburton, R. and Gall, G. A. E. 1979. Electrophoretic variation and differentiation in four strains of domesticated rainbow trout (*Salmo gairdneri*). Can. J. Genet. Cytol. **21**: 81-94.

Clayton, J. W. and Tretiak, D. N. 1972. Amino-citrate buffers for pH control in starch gel electrophoresis. J. Fish. Res. Board Can. **29**: 1169-1172.

Cross, T. F. and Payne, R. H. 1977. Nadm-isocitrate dehydrogenase polymorphism in the Atlantic salmon *Salmo salar*. J. Fish. Biol. **11**: 493-496.

Fuerst, P. A., Chakraborty, R and Nei, M. 1977. Statistical studies on protein polymorphism in natural population. I. Distribution of single locus heterozygosity. Genetics, **86**: 455-483.

Gold, J. R. 1977. Systematics of western North American trout (*Salmo*) with notes on the redband trout of Sheepheaven Creek, California. Can. J. Zool. **55**: 1858-1873.

Mac Crimmon, R. H. 1971. World distribution of rainbow trout (*Salmo gairdneri*) J. Fish. Res. Board Can. **28**: 663-704.

May, B., Wright, J. E. and Stoneking, M. 1979. Joint segregation of biochemical loci in salmonidae; Results from experiments with Salvelinus and review of the literature on other species. J. Fish. Res. Board Can. **36**: 1114-1128.

Nei, M. 1975. Molecular population genetics and evolution. North-Holland, Amsterdam and New York.

Ridgway, G. J., Sherburne, S. W. and Lewis, R. D. 1970. Polymorphisms in the esterases of Atlantic herring. Trans. Am. Fish. Ser. C, **99**: 147-151.

Roberts, F. L., Wohnus, J. F. and Ohno, S. 1969. Phosphoglucomutase polymorphism in the rainbow trout, Salmo gairdneri. Experientia, **25**: 1109-1110.

Schmidtke, J., Dunkhase, G. and Engel, W. 1975. Genetic variation of phosphoglucose isomerase isoenzymes in fish of the orders ostariophysi and isospondyli. Comp. Biochem. Physiol. **50B**: 395-398.

Utter, F. M. and Allendorf, F. W. 1977. Determination of the breeding structure of steelhead populations through gene frequency analysis. *Edited by* T. J. Hassler and R. R. Van Kirk. Calif. Coop. Fisheries Res. Unit, Spec. Report. pp. 1-77.

Utter, F. M., Hodgins, H. O., Allendorf, F. W., Johnson, A. G. and Mighell, J. L. 1973. Biochemical variants in Pacific salmon and rainbow trout; their inheritance and application in population studies. *In* Genetics and mutagenesis of fish. *Edited by* H. H. Schroder. Springer-Verlag, Berlin, Heidelberg, New York.

Wright, J. E., Heckman, J. R. and Atherton, L. M. 1975. Genetic and developmental analysis of LDH Isozymes in trout. *In* Isozymes, III, Developmental biology. *Edited by* C. L. Markert. Academic Press, New York.
<table><caption>APPENDIX TABLE I
Allelic frequencies for comparison of the 13 strains of domesticated rainbow trout</caption><thead><tr><th></th><th>S<sub>1</sub>-77</th><th>S<sub>1</sub>-78</th><th>S<sub>2</sub></th><th>S<sub>3</sub></th><th>Univ. of Washington</th><th>Chamber Creek</th><th>Mac Creary</th><th>West Virginia</th><th>RTH</th><th>RTHX</th><th>RTS</th><th>RTW</th><th>RTV</th></tr></thead><tbody><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Agpdh-2</td></tr><tr><td>α-gpdh (b)</td><td>0.0160</td><td>0.0</td><td>0.0</td><td>0.0250</td><td>0.0</td><td>0.0</td><td>0.1120</td><td>0.0</td><td>0.0020</td><td>0.0030</td><td>0.0</td><td>0.0090</td><td>0.2510</td></tr><tr><td>α-gpdh (a)</td><td>0.9840</td><td>1.000</td><td>1.0000</td><td>0.9750</td><td>1.0000</td><td>1.00000</td><td>0.8880</td><td>1.0000</td><td>0.9980</td><td>0.9970</td><td>1.0000</td><td>0.9910</td><td>0.7490</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Idh-3</td></tr><tr><td>Idh-3 (1.31)</td><td>0.1030</td><td>0.0870</td><td>0.1000</td><td>0.0930</td><td>0.0065</td><td>0.0</td><td>0.1000</td><td>0.0</td><td>0.0010</td><td>0.0</td><td>0.0240</td><td>0.3820</td><td>0.0</td></tr><tr><td>Idh-3 (1.00)</td><td>0.5950</td><td>0.5870</td><td>0.6460</td><td>0.6120</td><td>0.8125</td><td>0.7790</td><td>0.8735</td><td>0.8750</td><td>0.6990</td><td>0.7330</td><td>0.7440</td><td>0.3400</td><td>0.9260</td></tr><tr><td>Idh-3 (0.69)</td><td>0.0720</td><td>0.0760</td><td>0.1000</td><td>0.0510</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.1730</td><td>0.1580</td><td>0.1030</td><td>0.0750</td><td>0.0</td></tr><tr><td>Idh-3 (0.38)</td><td>0.2300</td><td>0.2500</td><td>0.1540</td><td>0.2440</td><td>0.1810</td><td>0.2210</td><td>0.0165</td><td>0.1250</td><td>0.1270</td><td>0.1080</td><td>0.1280</td><td>0.2030</td><td>0.0740</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Idh-4</td></tr><tr><td>Idh-4 (1.31)</td><td>0.1030</td><td>0.0870</td><td>0.1000</td><td>0.0930</td><td>0.0065</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0010</td><td>0.0</td><td>0.0240</td><td>0.3820</td><td>0.0</td></tr><tr><td>Idh-4 (1.00)</td><td>0.5950</td><td>0.5870</td><td>0.6460</td><td>0.6120</td><td>0.8125</td><td>0.7790</td><td>0.8735</td><td>0.0</td><td>0.6990</td><td>0.7330</td><td>0.7440</td><td>0.3400</td><td>0.9260</td></tr><tr><td>Idh-4 (0.69)</td><td>0.0720</td><td>0.0760</td><td>0.1000</td><td>0.0510</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.8750</td><td>0.1730</td><td>0.1580</td><td>0.1030</td><td>0.0750</td><td>0.0</td></tr><tr><td>Odh-4 (0.38)</td><td>0.2300</td><td>0.2500</td><td>0.1540</td><td>0.2440</td><td>0.1810</td><td>0.2210</td><td>0.0165</td><td>0.0</td><td>0.1270</td><td>0.1080</td><td>0.1280</td><td>0.2030</td><td>0.0740</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Mdh-3</td></tr><tr><td>Mdh-3 (1.25)</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0060</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0060</td><td>0.0</td></tr><tr><td>Mdh-3 (1.00)</td><td>0.7610</td><td>0.7630</td><td>0.9160</td><td>0.7950</td><td>0.9810</td><td>0.9530</td><td>0.9380</td><td>1.0000</td><td>0.9990</td><td>1.0000</td><td>0.9150</td><td>0.7520</td><td>1.0000</td></tr><tr><td>Mdh-3 (0.79)</td><td>0.2390</td><td>0.2370</td><td>0.0840</td><td>0.2050</td><td>0.0190</td><td>0.0470</td><td>0.0570</td><td>0.0</td><td>0.0010</td><td>0.0</td><td>0.0850</td><td>0.2420</td><td>0.0</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Mdh-4</td></tr><tr><td>Mdh-4 (1.25)</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0060</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0060</td><td>0.0</td></tr><tr><td>Mdh-4 (1.00)</td><td>0.7610</td><td>0.7630</td><td>0.9160</td><td>0.7950</td><td>0.9810</td><td>0.9530</td><td>0.9380</td><td>1.0000</td><td>0.9990</td><td>1.0000</td><td>0.9150</td><td>0.7520</td><td>1.0000</td></tr><tr><td>Mdh-4 (0.79)</td><td>0.2390</td><td>0.2370</td><td>0.0840</td><td>0.2050</td><td>0.0190</td><td>0.0470</td><td>0.0570</td><td>0.0</td><td>0.0010</td><td>0.0</td><td>0.0850</td><td>0.2420</td><td>0.0</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Pgm</td></tr><tr><td>Pgm (1.00)</td><td>0.7000</td><td>0.7370</td><td>0.9830</td><td>0.8100</td><td>1.0000</td><td>0.6390</td><td>0.8250</td><td>1.0000</td><td>0.6840</td><td>0.5610</td><td>0.9200</td><td>1.0000</td><td>0.9690</td></tr><tr><td>Pgm (0.70)</td><td>0.3000</td><td>0.2630</td><td>0.0170</td><td>0.1900</td><td>0.0</td><td>0.3610</td><td>0.1750</td><td>0.0</td><td>0.3160</td><td>0.4390</td><td>0.0800</td><td>0.0</td><td>0.0310</td></tr><tr><td colspan="14" style="text-align: center; font-weight: bold;">Locus Sod</td></tr><tr><td>Sod (1.61)</td><td>0.6600</td><td>0.7120</td><td>0.2800</td><td>0.6370</td><td>0.0</td><td>0.5580</td><td>0.5000</td><td>0.1560</td><td>1.000</td><td>0.9800</td><td>0.2950</td><td>0.1170</td><td>0.5570</td></tr><tr><td>Sod (1.00)</td><td>0.3400</td><td>0.2725</td><td>0.6400</td><td>0.3500</td><td>1.000</td><td>0.4420</td><td>0.5000</td><td>0.8440</td><td>0.0</td><td>0.0200</td><td>0.7050</td><td>0.8830</td><td>0.4420</td></tr><tr><td>Sod (0.39)</td><td>0.0</td><td>0.0130</td><td>0.0800</td><td>0.0130</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td><td>0.0</td></tr></tbody></table>