Analysis of a repetitive DNA family from Arabidopsis arenosa and relationships between Arabidopsis species

We have analysed a family of highly repetitive DNA from Arabidopsis arenosa (L.) Lawalrée [syn. Cardaminopsis arenosa (L.) Hayck] composed of AT-rich tandem repeats of 166–179 bp in head to tail organization. Sequence comparison between several repeat units revealed a high level of divergence of 4.5% to 25%. The sequence family shows more than 58% homology to satellite sequences described in Arabidopsis thallana (L.) Heynh. but no homology to other satellite repeats in the Cruciferae. Within the genus Arabidopsis the satellite sequence was found to be present in A. thaliana and Arabidopsis suecica (Fries) Norrlin, but not in Arabidopsis griffithiana (Boiss.) N. Busch and Arabidopsis pumila (Stephan) N. Busch. In situ hybridization to metaphase chromosomes of A. arenosa (2n=4x=32) showed the sequence to be localized at the centromeres of all 32 chromosomes with substantial hybridization along the chromosome arms. Using Southern hybridization and in situ hybridization, we give evidence that A. suecica is a hybrid of A. thaliana and A. arenosa. A considerable reorganization of the A. thaliana satellite sequence pAL1 occurred in the hybrid genome while no molecular change of the A. arenosa repeat was observed in the hybrid. Analysis of related repeats enabled differentiation between closely related genomes and are useful for the investigation of hybrid genomes.     

Studying genetics of adaptive variation in model organisms: flowering time variation in Arabidopsis lyrata

Arabidopsis thaliana has emerged as a model organism for plant developmental genetics, but it is also now being widely used for population genetic studies. Outcrossing relatives of A. thaliana are likely to provide suitable additional or alternative species for studies of evolutionary and population genetics. We have examined patterns of adaptive flowering time variation in the outcrossing, perennial A. lyrata. In addition, we examine the distribution of variation at marker genes in populations form North America and Europe. The probability of flowering in this species differs between southern and northern populations. Northern populations are much less likely to flower in short than in long days. A significant daylength by region interaction shows that the northern and southern populations respond differently to the daylength. The timing of flowering also differs between populations, and is made shorter by long days, and in some populations, by vernalization. North American and European populations show consistent genetic differentiation over microsatellite and isozyme loci and alcohol dehydrogenase sequences. Thus, the patterns of variation are quite different from those in A. thaliana, where flowering time differences show little relationship to latitude of origin and the genealogical trees of accessions vary depending on the genomic region studied. The genetic architecture of adaptation can be compared in these species with different life histories.     

Genetic diversity and invasibility: a test using a model system with a novel experimental design

Biological invasion is one aspect of ecosystem function that may be controlled by the biological diversity of the invaded community, and there have been a number of recent studies that investigated relationships between diversity and invasibility. Most experimental studies report that higher species or functional group diversity increases resistance to invasion, but the role of genetic diversity is unknown. We used a model organism, Arabidopsis thaliana (Brassicaceae), to investigate relationships between genotypic richness and community invasibility by creating communities with 1, 2, 4, and 8 genotypes of A. thaliana at constant low (417 plants m⁻²) and high (834 plants m⁻²) densities, that once established, were invaded with a congener, Arabidopsis suecica. To reduce the potential effects of methodological confounding related to “sampling effects,”“variance reduction effects,” or confounding of abundance with diversity, we (1) created random communities from a relatively large pool of functionally and phenotypically similar genotypes, (2) evaluated individual and community traits across richness treatments, and (3) analyzed similarity of communities within treatments (for “quasi- replication”) and between adjacent treatments (for “nestedness”). Genotypic richness had no effect on A. suecica demography (emergence, survivorship), size (biomass, rosette area), or reproductive potential (rates of bolting and fruiting or number and size of bolts). In contrast, the density of A. thaliana genotypes had strong effects on the size and reproductive potential of A. suecica, which suggests that characteristics of the recipient community other than genotypic richness (e.g. light) form the most important determinant of community invasibility. Individual- and community-level traits of community members (cover, biomass, survivorship) did not differ among richness treatments, and within- and between-treatment similarity was reduced (relative to other recent experiments) but not eliminated. We evaluate our results vis-a-vis recent analyses of diversity-invasibility experiments, and provide directions for future investigations of genetic diversity.     

CHLOROPLAST DNA AND ISOZYME DIVERSITY IN TWO MIMULUS SPECIES (SCROPHULARIACEAE) WITH CONTRASTING MATING SYSTEMS

Levels of cpDNA and isozyme diversity were contrasted between the mixed-mating M. guttatus and its highly selfing congener M. micranthus (Scrophulariaceae). Compared to M. micranthus, M. guttatus has two to four times higher diversity for both cpDNA and isozyme variation on a species-wide level. The selfing M. micranthus also has 1.5 to three times lower within-population allozyme variation and a greater proportion of its variation distributed among rather than within populations. Chloroplast DNA is here inferred to be uniparentally inherited. Thus the mating system has no effect on the transmission of the chloroplast genome. Since both cpDNA and isozyme variation are similarly reduced in M. micranthus, factors other than the mating system are hypothesized to be responsible for the observed decrease in species-wide genetic variation in M. micranthus. A recent origin of M. micranthus from a limited number of M. guttatus populations is suggested. Consequently, molecular variation is reduced in M. micranthus due to a bottleneck effect. These data generally demonstrate that levels of cpDNA variation may be high enough in some species to infer evolutionary processes below the species level.     

The effect of the nanoflagellateTetraselmis suecica on the growth and survival of grunion,Leuresthes tenuis,larvae

Synopsis The nanoflagellateTetraselmis suecica was tested both as the sole food source and as a diet complement toArtemia nauplii for grunion,Leuresthes tenuis, larvae. A total of 4800 grunion larvae, obtained through artificial insemination and incubation, were cultivated under laboratory conditions. Growth and survival rates were registered for 14 days in two experimental series. In the first series the nanoflagellateT. suecica was offered as the sole food source at five different concentrations. Survival and growth increased in agreement with the increase inT. suecica concentration. In the second series,Artemia nauplii were offered at six concentration levels. This series was divided into two groups: the nanoflagellateT. suecica was added to one group at a concentration of 5000 cells ml^–1; the other group was maintained without nanoflagellates. In this series, survival and growth were directly related to nauplii concentration, but significant effects of the nanoflagellates were evident only in relation to the survival; the greatest difference (58% without nanoflagellates vs. 69% with nanoflagellates) was observed at anArtemia concentration of 1000 nauplii 1^–1. The mechanism responsible for increased survival ofL. tenuis larvae in presence of phytoplankton is unclear.     

Changes in the isozyme composition of antioxidant enzymes in response to aminotriazole in leaves ofArabidopsis thaliana

The changes in isozyme profiles of catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX), and glutathione reductase (GR) during severe deactivation of total CAT activity by aminotriazole (AT) treatment were investigated in the leaves ofArabidopsis thaliana (Columbia ecotype) in relation to H₂O₂-mediated oxidative stress. In spite of striking deactivation of total CAT activity by 0.1 mM AT, there were no significant differences in H₂O₂ levels or total leaf soluble protein contents including a Rubisco in both the control and AT-treated leaves. On the other hand, one specific protein band (molecular mass, 66 kD) was observed on the SDS-gel from leaf soluble proteins whose staining intensity was strikingly enhanced by AT treatment for 6 h. However, this band disappeared at 12 h. In the native-gel assays of CAT, POD, APX and GR isozymes, AT remarkably inhibited the expression of the CAT1 isozyme with no effects on CAT2 and CAT3, and generally had no effect on POD isozyme profiles. However, AT stimulated the intensity of activities of pre-existing APX1 and GR1 isozymes. In particular, it induced a new synthesis of one GR isozyme. Therefore, these results collectively suggest that a striking deactivation of total CAT activity by AT inA. thaliana leaves largely results from the suppression of CAT1 isozyme, and that APX1, GR1, and a newly synthesized GR isozyme could complement the role of CAT1 to metabolize H₂O₂ into non-toxic water.     

Induction of telomere-mediated chromosomal truncation and stability of truncated chromosomes in Arabidopsis thaliana

Minichromosomes possess functional centromeres and telomeres and thus should be stably inherited. They offer an enormous opportunity to plant biotechnology as they have the potential to simultaneously transfer and stably express multiple genes. Segregating independently of host chromosomes, they provide a platform for accelerating plant breeding. Following a top‐down approach, we truncated endogenous chromosomes in Arabidopsis thaliana by Agrobacterium‐mediated transfer of T‐DNA constructs containing telomere sequences. Blocks of A. thaliana telomeric repeats were inserted into a binary vector suitable for stable transformation. After transfer of these constructs into the natural tetraploid A. thaliana accession Wa‐1, chromosome truncation by T‐DNA‐induced de novo formation of telomeres could be confirmed by DNA gel blot analysis, PCR (polymerase chain reaction), and fluorescence in situ hybridisation. The addition of new telomere repeats in this process could start alternatively from within the T‐DNA‐derived telomere repeats or from adjacent sequences close to the right border of the T‐DNA. Truncated chromosomes were transmissible in sexual reproduction, but were inherited at rates lower than expected according to Mendelian rules.     

The influence of matrix attachment regions on transgene expression in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> wild type and gene silencing mutants

Many studies in both animal and plant systems have shown that matrix attachment regions (MARs) can increase the expression of flanking transgenes. However, our previous studies revealed no effect of the chicken lysozyme MARs (chiMARs) on transgene expression in the first generation transgenic Arabidopsis thaliana plants transformed with a β-glucuronidase gene (uidA) unless gene silencing mutants were used as genetic background for transformation. In the present study, we investigated why chiMARs do not influence transgene expression in transgenic wild-type Arabidopsis plants. We first studied the effect of chiMARs on transgene expression in the progeny of primary transformants harboring chiMAR-flanked T-DNAs. Our data indicate that chiMARs do not affect transgene expression in consecutive generations of wild-type A. thaliana plants. Next, we examined whether these observed results in A. thaliana transformants are influenced by the applied transformation method. The results from in vitro transformed A. thaliana plants are in accordance with those from in planta transformed A. thaliana plants and again reveal no influence of chiMARs on transgene expression in A. thaliana wild-type transformants. The effect of chiMARs on transgene expression is also examined in in vitro transformed Nicotiana tabacum plants, but as for A. thaliana, the transgene expression in tobacco transformants is not altered by the presence of chiMARs. Taken together, our results show that the applied method or the plant species used for transformation does not influence whether and how chiMARs have an effect on transgene expression. Finally, we studied the effect of MARs (tabMARs) of plant origin (tobacco) on the transgene expression in A. thaliana wild-type plants and suppressed gene silencing (sgs2) mutants. Our results clearly show that similar to chiMARs, the tobacco-derived MARs do not enhance transgene expression in a wild-type background but can be used to enhance transgene expression in a mutant impaired in gene silencing. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users. Miguel F.C. De Bolle, Katleen M.J. Butaye Contributed equally to this work     

Source verification of mis-identified Arabidopsis thaliana accessions

A major strength of Arabidopsis thaliana as a model lies in the availability of a large number of naturally occurring inbred lines. Recent studies of A. thaliana population structure, using thousands of accessions from stock center and natural collections, have revealed a robust pattern of isolation by distance at several spatial scales, such that genetically identical individuals are generally found close to each other. However, some individual accessions deviate from this pattern. While some of these may be the products of rare long‐distance dispersal events, many deviations may be the result of mis‐identification, in the sense that the data regarding location of origin data are incorrect. Here, we aim to identify such discrepancies. Of the 5965 accessions examined, we conclude that 286 deserve special attention as being potentially mis‐identified. We describe these suspicious accessions and their possible origins, and advise caution with regard to their use in experiments in which accurate information on geographic origin is important. Finally, we discuss possibilities for maintaining the integrity of stock lines.     

The chromosome morphology ofArabidopsis thaliana (L.)Heynh. and some remarks on the problem ofHylandra suecica (Fr.) Löve

The chromosome number ofA. thaliana from three localities in Central Bohemia was found to be 2n=10. All the chromosomes (length 1,5–2,6μm) belong to the atelocentric type, four pairs (m) having the centromere in the median and one pair (sm) in the submedian region. In connection with the discussion on the origin ofH. suecica the author presents the following preliminary results: a) the failure to cross the tetraploidCardaminopsis arenosa (L.)Hayek withA. thaliana; b) the successful crossing of the diploidC. petraea (L.)Hiit. withA. thaliana; c) the discovery of a diploid population ofC. arenosa (2n=16) in the Tatra Mts. (Czechoslovakia).     

Phenotypic responses to light,water, and nutrient conditions in the allopolyploid Arabidopsis suecica and its parent species A. thaliana and A. arenosa: Does the allopolyploid outrange its parents?

Polyploid species possess more than two sets of chromosomes and may show high gene redundancy, hybrid vigor, and masking of deleterious alleles compared to their parent species. Following this, it is hypothesized that this makes them better at adapting to novel environments than their parent species, possibly due to phenotypic plasticity. The allopolyploid Arabidopsis suecica and its parent species A. arenosa and A. thaliana were chosen as a model system to investigate relationships between phenotypic plasticity, fitness, and genetic variation. Particularly, we test if A. suecica is more plastic, show higher genetic diversity, and/or have higher fitness than its parent species. Wild Norwegian populations of each species were analyzed for phenotypic responses to differences in availability of nutrient, water, and light, while genetic diversity was assessed through analysis of AFLP markers. Arabidopsis arenosa showed a higher level of phenotypic plasticity and higher levels of genetic diversity than the two other species, probably related to its outbreeding reproduction strategy. Furthermore, a general positive relationship between genetic diversity and phenotypic plasticity was found. Low genetic diversity was found in the inbreeding A. thaliana. Geographic spacing of populations might explain the clear genetic structure in A. arenosa, while the lack of structure in A. suecica could be due to coherent populations. Fitness measured as allocation of resources to reproduction, pointed toward A. arenosa having lower fitness under poor environmental conditions. Arabidopsis suecica, on the other hand, showed tendencies toward keeping up fitness under different environmental conditions.     

Analysis of remote asymmetric somatic hybrids between common wheat and <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Callus-derived protoplasts of common wheat (Triticum aestivum L. cv. Hesheng 3) irradiated with ultraviolet light were fused by using the PEG method with cell suspension-derived protoplasts of Arabidopsis thaliana. Regenerated calli and green plants resembling that of wheat were obtained. The hybrid nature of putative calli and plants were confirmed by isozyme, random amplified polymorphic DNA and genomic in situ hybridization (GISH) analyses. GISH results indicated that 1∼3 small chromosome fragments of A. thaliana were found introgression into the terminals of wheat chromosomes, forming highly asymmetric hybrids. Cytoplasmic genome tests did not show any cytoplasmic genetic materials from A. thaliana. However, variations from the normal wheat cytoplasmic genome were found, indicating recombination or rearrangement occurred during the process of somatic hybridization. The chromosome elimination in the asymmetric somatic hybridization of remote phylogenetic relationship was discussed. A miniature inverted-repeat transposable element related sequence was found by chance in the hybrids which might accompany and impact the process of somatic hybridization. Jingyao Deng and Haifeng Cui provided same contribution to this work.     

Genetics of allozyme variation in Gossypium arboreum L. and Gossypium herbaceum L. (Malvaceae)

Summary Seed protein extracts from 90 accessions of Gossypium arboreum and 70 accessions of Gossypium herbaceum were electrophoretically analyzed for isozyme variation. Eighteen enzyme systems were resolved, ten of which were polymorphic among accessions. No within accession isozyme variation was observed within these highly inbred lines. A minimum of 24 genes encode the isozymes resolved and data is presented for codominant inheritance at 13 loci. Tests for non-random joint segregation in 63 of the 78 possible two-locus combinations from the 13 characterized loci give evidence for four pairs of linked genes (Lap2/Me1 [r=0.160+/-0.027], Lap2/Pgi1 [r= 0.285+/-0.055], Mdh6/Tpi1 [r= 0.197+/-0.028], and 6Pgd2/6Pgd3[r 0.000]. Numerous presumptive duplicate isozyme loci were observed and these were usually expressed as patterns of nonsegregating heteromultimers within accessions. Single gene expression was also observed at several loci. The observed results are in agreement with those of previous cytological investigations which have proposed the polyploid origin of the diploid Old World Gossypiums.     

Effects of salicylic acid on paraquat tolerance in<Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> plants

Foliar spraying ofArabidopsis thaliana (Columbia ecotype) plants with a 1.0-mM salicylic acid (SA) solution significantly improved their tolerance to subsequent paraquat (PQ)-induced oxidative damage. Leaf injuries, including losses of chlorophyll, protein, and fresh weight, were reduced. Our analysis of antioxidant enzymes in the leaves showed that SA pre-treatment effectively retarded rapid decreases in the activities of Superoxide dismutase (SOD), catalase, and ascorbate peroxidase that are normally associated with PQ exposure. In addition, guaiacol peroxidase activity was remarkably increased. In a native gel assay of peroxidase (POD) isozymes, staining activity of the POD1 isozyme, which disappeared in plants exposed only to 10 μM PQ, was significantly recovered by the 1.0-mM SA pre-treatment POD2 isozyme activity was also pronounced in all SA-treated plants compared with the control. A 12-h SA pre-treatment, without subsequent PQ stress, also caused a small increase in the endogenous H₂O₂ content that accompanies the symptoms of mild leaf injuries. This enhanced level occurred in parallel with a slight SOD increase and a catalase decrease. From our results, it can be assumed that, due to the small increase in SOD as well as catalase inactivation via SA pre-treatment, a moderate increase in H₂O₂ levels may occur. In turn, a large induction of guaiacol peroxidase leads to enhanced PQ tolerance inA. thaliana plants.     

Circumscription of species in the genus Sirodotia (Batrachospermales,Rhodophyta) based on molecular and morphological data

Species level taxonomy and phylogeographical distribution patterns in the freshwater rhodophyte Sirodotia are resolved through phylogenetic inferences based on rbcL and cox2–3 sequence data. Previous studies focused on the taxonomy of specific Sirodotia species or the distributions across a limited geographical region. Our molecular phylogenies included samples attributable to five recognized Sirodotia species and include collections from Australia, Brazil, Costa Rica, Canada, Finland, Mexico, New Zealand, South Africa and the United States. Both rbcL and cox2–3 phylogenies inferred S. suecica, S. tenuissima and S. goebelii as a monophyletic group with little sequence divergence. This result supports the synonymy of S. tenuissima and S. goebelii with S. suecica (the species name with priority). Within this clade, samples collected from Australia and New Zealand formed a monophyletic group with no other discernible phylogeographical patterns within S. suecica. This result seems to be somewhat unusual in the Batrachospermales, as other species have shown greater genetic variation among geographically distant locations. As in previous studies, S. huillensis and S. delicatula were inferred as a separate species based on the rbcL phylogeny, supporting the current taxonomy. A specimen of S. aff. huillensis from South Africa, may represent a new species but further research is necessary before it can be designated as such.     

Genetic Diversity in Natural Populations of Arabidopsis thaliana (L.) Heynh. from Karelia

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 (P _99% = 43.7, H _obs = 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 (G _ST = 0.583).     

Oil production from five marine microalgae for the production of biodiesel

Marine microalgae were studied as potential resources for the production of biodiesel. Five marine microalgae, Tetraselmis suecica, Phaeodactylum tricornutum, Chaetoceros calcitrans, Isochrysis galbana, and Nannochloropsis oculata were cultured in f/2 media, 12:12 L:D cycle at 20 ± 1°C with a light intensity of 36.3 μmol/m²/sec using a 15-L circular cylindrical photobioreactor. The dry cell weight, specific growth rate, biomass productivity, oil content and fatty acid composition of palmitic acid, stearic acid, oleic acid, linoleic acid, and linolenic acid of microalgae were determined. T. suecica, I. galbana, and N. oculata showed high dry cell weights of 0.58, 0.57, and 0.57 g/L, respectively. The culture period of T. suecica to reach the stationary phase was 9 days. On the other hand, N. oculata showed the longest culture period of 28 days to reach the stationary phase. T. suecica absorbed nitrate at the initial stages of cell growth, decreasing the nitrate concentration to 0.5 mg/L on day-7 of the culture. The highest oil contents were observed in P. tricornutum with a 25.31% dry cell weight and I. galbana with a 23.15% dry cell weight on day-9 after the stationary phase. I. galbana showed 417.33 mg of palmitic acid per g oil and T. suecica showed 235.61 mg of oleic acid per g oil. Stearic acid, linoleic acid, and linolenic acid did not exceed 30.02 mg/g oil in any of the microalgae. T. suecica showed the shortest culture period of 9 days to reach the stationary phase. Therefore, the highest biomass production of 0.58 g/L was obtained and I. galbana showed high biomass production of 0.57 g/ L and oil content of 23.15% of dry cell weight. Therefore, T. suecica and I. galbana can be selected as a potential candidate for the production of biodiesel.     

Retina-specific LDH isozyme in blueback herring, Alosa aestivalis (Mitchell), and alewife, Aiosa pseudoharengus (Wilson)

The retina of 390 Alosa aestivalis and 410 Alosa pseudoharengus have been examined by means of starch-gel electrophoresis. The retina-specific E₄ isozyme has been found to occur in all the fish examined. This study demonstrates for the first time that the E₄ isozyme occurs in A. aestivalis. Because the E₄ isozyme is not polymorphic and has an identical mobility in A. pseudoharengus and in A. aestivalis it is neither suitable for use as a species identification characteristic nor a population marker. Alosa aestivalis and Alosa pseudoharengus are two commercially important ana-dromous species of fish in New Brunswick, Canada. These two species may occur together in the same spawning runs but w^rhile A. pseudoharengus has a wide distribution along the East coast of North America (Leim & Scott, 1966) A. aestivalis occurs in a very limited area in Canada where it is at the northern limit of its range and because of increasing threat of pollution has been listed by McAllister (1970) as one of 17 endangered species. These two species of fish are morphologically very similar and can only be separated by the colour of the abdominal peritoneum (Leim & Scott, 1966). McKenzie (1973) has compared these two species by means of protein electrophoresis and found that while the muscle myogen patterns were species specific, the LDH patterns were the same in both species. He described these two species as five isozyme fish showing the LDH isozymes A₄, A₃B, A₂B₂, AB₃ and B₄. Since Horowitz & Whitt (1972) have reported the presence of the E₄ isozyme in the retina of some teleosts including. A. pseudoharengus but not including A. aestivalis, I considered it worth-while to re-examine A. aestivalis and A. pseudoharengus to find out whether A. aestivalis possessed this isozyme and, if so, whether the mobility of the isozyme could be used as a species identification characteristic and as a population marker. The fish used in this study were collected from five different locations during the 1971 spring migration period and held deep frozen for eight months before they were examined. They were identical to the specimens used for the study reported by McKenzie (1973) where the collection dates, numbers of fish and geographic locations are given. One eyeball from each of 390 A. aestivalis and 410 A. pseudoharengus was removed. Each eyeball was homogenized in 10 drops of distilled water and allowed to stand for one hour at 4^oC. The samples were then centrifuged for 10 min at 12 000 g. The supernatants were used immediately for vertical starch-gel electrophoresis. The apparatus used was that described by Boyer & Hiner (1963). The conditions of electrophoresis were the same as used by Saunders & McKenzie (1971). The LDH bands were stained by the deposition of blue formazan dyes in the regions of LDH activity. The stain formula and details of methods are given in Whitt (1970). The LDH isozyme patterns of all the fish examined were identical. The location of the isozymes A₄, A₃B, A₂B₂, AB₃ and B₄ are indicated in Fig. 1. A₄ is abundant in muscle while B₄ is abundant in heart. Because the LDH subunits assemble preferentially into homodimeric pairs before forming tetramers (Markert & Ursprung, 1971). A₄. A₂B₂ and B₄ show up in electropherograms as strong bands while A₃B and AB₃ show up as weak bands. The retina-specific isozyme E₄ is shown between B₄ and AB₃. Whitt (1970) has already demonstrated that A. pseudoharengus is a five isozyme fish. This has been confirmed by McKenzie (1973) who also compared both A. pseudoharengus and A. aestivalis and found these five isozymes had identical mobilities in both species of fish. The present study demonstrates for the first time that the retina-specific E₄ occurs in A. aestivalis where it has the same electro-phoretic mobility as that of A. pseudoharengus. The patterns are the same and do not appear to vary with geographic origin of the fish. The reason why the presence of the E₄ isozyme was demonstrated in the present study and not in the previous one (McKenzie, 1973) is because of the method used. In that study the migration length was not long enough to sufficiently separate the enzymes. In the present study vertical starch-gel electrophoresis which allows for long migration distances was used. As has already been shown for the A-B isozymes (McKenzie, 1973), the E₄ isozyme is not polymorphic in these two species of fish. It therefore has no use as apopulation marker. Because the E₄ isozyme has an identical mobility in both species of fish, it cannot be used as a species identification characteristic.     

DISTRIBUTION AND SYSTEMATICS OF THE FRESHWATER GENUS SIRODOTIA(BATRACHOSPERMALES,RHODOPHYTA) IN NORTH AMERICA1

Multivariate morphometrics and image analysis were used to determine the number of well-delineated infrageneric taxa of Sirodotia in North America. Three groupings were distinguished from 25 populations examined from Newfoundland and Quebec in the north to central Mexico in the south. These groupings were statistically related to 10 type specimens, and the following species were recognized: Sirodotia huillensis (Welwitsch ex W. et G. S. West) Skuja (syn. S. ateleia Skuja), S. suecica Kylin (syn. S. acuminata Skuja ex Flint and S. fennica Skuja), and S. tenuissima (Collins) Skuja ex Flint. These species are differentiated on the basis of whorl shape and degree of separation at maturity (S. suecica, rounded and appressed; S. huillensis and S. tenuissima, truncated apex and separated), the density of spermatangia (S. huillensis, dense clusters; S. suecica and S. tenuissima, sparsely aggregated), and the mode of germination of the gonimoblast initial (S. suecica and S. tenuissima, from the nonprotuberant side of the fertilized carpogonium; S. huillensis from the protuberant side). Sirodotia huillensis was found only in the desert-chaparral, whereas S. suecica and S. tenuissima occurred from south-temperate to boreal regions in cool (temperature 8–18° C), low ion (specific conductance 10–99 μS · cm^?1), and mildly acidic to neutral (pH 5.7–7.3) waters.