Use of recombinant substitution lines in the construction of RFLP-based genetic maps of chromosomes 6A and 6B of tetraploid wheat (Triticum turgidum L.)

RFLP-based genetic maps of chromosomes 6A and 6B of Triticum turgidum have been constructed using data obtained by the study of Triticum turgidum var durum cv Langdon-T. t. var dicoccoides recombinant substitution lines (RSLs) supplemented with data obtained from F₃ families derived from Langdon dicoccoides 6A and 6B disomic substitution lines. The average RFLP frequencies detected for the two chromosomes in a test of 45 DNA clones with six restriction enzymes were 56% and 53%, respectively, and a subset of 32 clones gave frequencies of 75% and 72%, respectively. Seventeen loci were mapped in 6A and 18 in 6B. With the possible exception of 5 loci in the centromeric region of 6A, all of the mapped 6A and 6B loci are located in the same arm as are homologous loci in hexaploid wheat, and the linear order of the loci is the same in the two chromosomes, except possibly close to the centromere. Major differences in genetic distances exist between homologous loci located in the proximal regions of the 6AL and 6BL linkage groups, however, the distances being much larger in the former than in the latter. The 6B maps that were constructed using data from both the RSL and the F₂ populations and using data from the RSL population alone closely resemble one another, indicating that the 6B RSL population, composed of 85 lines, can be reliably used for genetic mapping. Additional studies must be conducted before the utility of the 6A RSL population, composed of 66 lines, can be adequately assessed.     

Temporal variation of Drosophila melanogaster Adh allele frequencies,inversion freqencies,and population sizes

The frequencies of cosmopolitan inversions and of Adh alleles in D. melanogaster, and population sizes of D. melanogaster and D. simulans have been monitored for two years in seven south-eastern Queensland sites. D. melanogaster population sizes showed greater summer increases and winter decreases than those of D. simulans. Adh allele frequencies showed no seasonal or yearly variation, and no significant correlations with environmental variables after accounting for linkage disequilibrium with In(2L)t. Some inversion frequencies showed regular summer to winter declines, yearly variation, and significant correlations with population size. Thus, the temporal variation of some inversion frequencies, unlike that for Adh allele frequencies, paralleled previously reported frequency variation over latitude.     

Deletion of the phosphoglucose isomerase structural gene makes growth and sporulation glucose dependent in Saccharomyces cerevisiae

Summary The structural gene PG11 coding for phosphoglucose isomerase was replaced by the LEU2 gene in the genome of Saccharomyces cerevisiae. Plasmids carrying the LEU2 gene between genomic regions flanking the PG11 gene were constructed and used to transform a PGI1/pgi1 diploid strain. Stable transformants lacking the PGI1 allele were isolated. Southern analysis of their meiotic products showed that haploid strains with a deletion of 1.6 kb within the 2.2 kb PG11 coding region were viable. Thus, the PGI1 gene is not essential in yeasts. However, unlike pgi1 mutants with residual phosphoglucose isomerase activity, no growth was detected in the pgi1 haploid strains when fructose was supplied as sole carbon source. The wild-type growth rate could be restored by adding 0.1% glucose to the medium. Furthermore, pgi1 mutants with residual enzymatic activity grew very slowly on fructose-supplemented media containing up to 2% glucose. Strains carrying the deletion allele, however, failed to grow at glucose concentrations higher than 0.5%. Also the pgi1 strains did not grow in glucose as sole carbon source. On the other hand pgi1/pgi1 diploid strains did not sporulate on the usual acetate medium. This defect could be alleviated by the addition of 0.05% glucose to the sporulation medium. Under these conditions the pgi1 mutants sporulated with an efficiency of 25% compared with the wild type. These results suggest that (a) the phosphoglucose isomerase reaction is the only step catalysing the interconversion of glucose-6-P and fructose-6-P, (b) glucose-6-P is essential in yeasts, and (c) the oxidation of glucose-6-P through the glucose-6-P dehydrogenase reaction is not sufficient to support growth in yeasts.     

Influence of parthenogenetic reproduction on the genotypic constitution and evolutionary success of populations and species

The life cycle of Daphnia commonly includes parthenogenesis, sexual reproduction, and diapause. Although there are no genotypic transformations during diapause, it separates the clonal and pseudopanmictic states of the population. During parthenogenetic reproduction primarily polymorphic natural populations gradually degenerate into a mixture of a few clones. As resting eggs are usually produced sexually (= by means of sexual reproduction), after the diapause the vast diversity of individual genotypes normal for panmictic populations is observed. It is the competitive interactions between parthenogenetic clones which again eventually decrease the genotypic polymorphism. Forms in which the sexual process and diapause are rare (or species which are represented by parthenogenetic populations in one part of their area and by bisexual ones in another) demonstrate the most significant differences between clonal and panmictic populations.Parthenogenesis, apomixis and other kinds of reproduction without genetic recombination are widely spread in branchiopod crustaceans (Notostraca, Anostraca, Conchostraca, Cladocera) and in many other animals and plants. Moreover, reproduction without recombination plays an important role in evolution of faunas and floras. It is emphasized that non-recombinating races and species having high heterozygosity but a low level of genetic variation, enjoy short term advantages, but die out after change in the environment.     

The human apolipoprotein B 3′ hypervariable region: detection of eight new alleles and comparisons of allele frequencies in blacks and whites

We investigated common length polymorphisms in the hypervariable region located 3 to the human gene encoding apolipoprotein B (APOB 3 HVR) as part of the Pathobiological Determinants of Atherosclerosis in Youth (PDAY) study. PDAY is a multicenter study of young persons who died of external causes (accident, homicide, and suicide). The APOB 3 HVR contains multiple copies of AT-rich tandem repeats (15bp) called hypervariable elements (HVE). Using polymerase chain reaction (PCR) to amplify APOB sequences in hepatic DNA samples, we identified 22 different HVR alleles among 232 PDAY cases. In addition to 14 previously identified alleles, we detected 8 new alleles that had not been observed in population surveys. Of these new alleles, 7 were present only in black cases. We also examined distributions of HVR allele frequencies for blacks and whites. The frequency distributions for whites did not differ from those from previous studies of French populations (P=0.3811) and Austrian populations (P= 0.1885). In contrast, the allele frequency distribution for blacks differed from whites (P<0.001). Blacks had higher frequencies of smaller alleles (33 repeats) and larger alleles (37 repeats) than whites. We also sequenced specific HVR alleles to identify differences responsible for size variation. The most frequent alleles were identical in sequence to HVR alleles described in previous studies. However, one allele was not identical in sequence to an equivalent-sized allele from a previous study. In all likelihood, detection of sequence substitutions in the APOB 3 HVR would result in an even greater amount of allelic variability than detected by size differences alone.     

On morphological variation in Keratella cochlearis populations from Holstein lakes (Northern Germany)

Keratella cochlearis occurs in many Holstein lakes (northern Germany) as three well defined and separated forms: cochlearis, hispida, and tecta, each showing very little variation between the lakes. The present data show that the tecta form did not originate from a Lauterborn cycle.     

Geographic structuring of molecular and morphological polymorphism in Pyrenean populations of the snail Cepaea nemoralis

To determine whether geographic patterns of variation in the frequency of shell polymorphisms in the land snail Cepaea nemoralis mark regions of extensive genetic differentiation, allele frequencies at six polymorphic enzyme-encoding loci were analyzed electrophoretically in 74 samples from the central part of the Spanish Pyrenees. Within a large area effect for unbanded shells on the southern slope of the Pyrenees, there is no enzyme locus at which allele frequencies are homogeneous. Because this morphological area effect does not mark a region of pervasive genetic change, the genetic structure of populations in the region studied does not conform to recent models of area effect speciation. Factor analysis of allele frequencies at three loci controlling shell polymorphisms and six enzyme loci separated populations on the southern slope into two groups, one of which is similar to the group of populations on the northern slope. There is therefore considerable geographic structuring of both molecular and morphological polymorphism in this species.     

Microsatellite markers in the hexaploid Prunus domestica species and parentage lineage of three European plum cultivars using nuclear and chloroplast simple-sequence repeats

In a previous study, cDNA microsatellite markers were described in apricot (Prunus armeniaca L.). Specific PCR primers were designed to amplify the microsatellite-containing regions from genomic DNA in different Prunus species. In the present work, cDNA microsatellite markers were developed in the hexaploid Prunus domestica L. species and polymorphism was ascertained in a segregating plum population. Co-dominant mendelian segregation of alleles was demonstrated and microsatellite polymorphism displayed up to 6 alleles per SSR locus per individual. Parentage lineage of three full-sib European plum cultivars (cv. Cacanska najbolja, Cacanska rana and Cacanska lepotica) was reconstructed by the analysis of the above nuclear SSR markers, completed by four chloroplastic microsatellite loci. The six most informative nuclear loci enabled discrimination between the three Cacak cultivars and unrelated individuals as well as the previously proposed parents, Wangenheim and Pozegaca. Data obtained support previous evidence that these cultivars originated from the Stanley cultivar. However, SSR analysis finally excluded Wangenheim as the other possible parent. Based on the results obtained with nuclear and chloroplast SSR loci, we propose the origin of those three Cacak cultivars in a cross between Stanley as the mother plant and Ruth gerstetter as the pollinator. Furthermore, we demonstrate the utility of these apricot SSR markers for genotype fingerprinting of the hexaploid plum cultivars.     

A simple method for maintaining high multiplication of Narcissus shoot cultures in vitro

A simple method for stimulating and maintaining high in vitro multiplication of Narcissus shoot clump cultures was developed. Shoot clumps were subjected either to normal cutting where leaves were trimmed to 20 mm in length at the beginning of each culture passage or to severe cutting where shoot clumps were cut down to the basal plate region removing all green tissue. Severe cutting at the beginning of each culture passage initially doubled the leaf multiplication, compared to normal cutting, but the difference between cutting treatments declined in successive passages. The improvement in leaf multiplication was maintained when shoot clumps were subjected to severe cutting only at every other culture passage, with no cutting in the alternate recovery passages. In vitro multiplication was increased by severe cutting in all seven Narcissus cultivars which were tested.Abbreviations NAA-1 naphthylacetic acid - BAP benzylaminopurine     

Molecular mapping of a new gene Rrs4 CI 11549 for resistance to barley scald (Rhynchosporium secalis)

Doubled haploid (DH) progeny from a cross between the scald susceptible barley (Hordeum vulgare L.) cultivar Ingrid and the resistant accession CI 11549 (Nigrinudum) was evaluated for resistance in the pathogen Rhynchosporium secalis (Oudem) J.J. Davis. Two linked and incompletely dominant loci confer resistance CI 11549 against isolate 4004. One is an allele at the complex Rrs1 locus on chromosome 3H close to the centromere; the other is located 22 cM distally on the long arm. The latter locus is designated Rrs4. In BC₃-lines into Ingrid from CI 2222 (another Nigrinudum) resistance seems governed by one locus close to the telomeric region of chromosome 7H, probably allelic to Rrs2. In neither case did we find any trace of the recessive gene rh8 reported to be present in Nigrinudum. Various resistance donors of Ethiopian origin designated as Nigrinudum, Jet or Abyssinian were identical to a great extent with respect to markers, but differed in resistance to different isolates of scald or in barley yellow dwarf virus (BYDV) resistance. The implications for their use as differentials in scald tests and screening of germplasm collections are discussed.     

Identification and confirmation of three neutral alleles conferring wide compatibility in inter-subspecific hybrids of rice (Oryza sativa L.) using near-isogenic lines

Wide-compatibility varieties (WCVs) are a special class of rice germplasm that is able to produce fertile hybrids when crossed to both indica and japonica subspecies. Previous studies determined Dular and 02428 as two WCVs and identified a number of QTLs as having large effects on fertility of inter-subspecific hybrids. In this study, we developed five near-isogenic lines (NILs) for three of the QTLs, f5, f6 and S5, by backcrossing and marker-assisted selection, using Dular and 02428 as the donors and Zhenshan 97 as the recipient. Three of the NILs each carried one introgressed allele, and two NILs each carried two introgressed alleles in combinations. The NILs were testcrossed to an indica tester Nanjing 11 and a japonica tester Balilla. The results showed that the f5 allele from Dular (f5-Du) is a neutral allele conferring wide compatibility, with a large effect on both pollen and spikelet fertility, and the f6 allele from Dular (f6-Du) is a neutral allele for spikelet fertility with smaller effect. The S5 allele from 02428 (S5-08) was confirmed to be a neutral allele for spikelet fertility. It is likely that f6 and S5 are the same locus as deduced by their genomic locations and effects. The results also showed that even in combination, two neutral alleles of different loci were not able to produce normal fertility hybrids in typical indica–japonica crosses. The implications of the findings in rice breeding programs are discussed.     

Mapping strategy for resistance genes in tomato based on RFLPs between cultivars: Cf9 (resistance to Cladosporium fulvum) on chromosome 1

Summary The contribution of introgressed regions derived from wild species to the genetic variation within the species of Lycopersicon esculentum was investigated by comparing the RFLP patterns of 2 introgression-free, obsolete cultivars (Moneymaker and Premier) and a modern cultivar (Sonatine) that carries at least 5 introgressed resistance genes. In this analysis 195 mapped nuclear markers were used in combination with 6 restriction enzymes. Among the 1170 probe-enzyme combinations tested, only 3 showed a polymorphism between the 2 introgression-free cultivars. On the other hand 24 probe-enzyme combinations were found to exhibit polymorphisms between Moneymaker and Sonatine. These represented ten polymorphic loci distributed among 5 linkage groups on chromosomes 1, 3, 4, 6, and 9.On the assumption that most of the polymorphic loci corresponded to introgressed chromosome segments of wild species carrying resistance genes, linkages between these loci and the component resistance genes were examined by RFLP analysis of pairs of near-isogenic lines differing only for one particular resistance gene, and a variety of commercial cultivars having different resistance gene compositions. Two of the polymorphic linkage groups could thus be ascribed to resistance genes whose map positions were already known: Cf2 on chromosome 6 and Tm2a on chromosome 9, whereas another marker, TG301 on chromosome 1, could be assigned to the Cladosporium fulvum resistance gene Cf9 with a hitherto disputable map position. By linkage analysis of a segregating F₂ population the genetic distance between the Cf9 gene and the marker TG301 was estimated at 5.5 ± 2.3 cM.     

Phylogenetic relationship of zeins and coixins as determined by immunological cross-reactivity and Southern blot analysis

Zeins from Zea mays L cv. Maya and coixins from Coix lacryma-jobi L. cv. Adlay were fractionated to obtain -, -, and -zein and -, -, and -coixin. The -coixins were composed of 4 polypeptide classes of 27 kDa (C1), 25 kDa (C2), 17 kDa (C4) and 15 kDa (C5) with solubility properties very similar to those of the 22 kDa and 19 kDa -zeins. Like the -zeins, the C1 and C2 -coixins corresponded to 80% of total Coix prolamins. The fraction corresponding to -coixin contained only one protein band of 22 kDa (C3). This coixin fraction has solubility properties similar to those of -zein and represents 15% of the total coixin. The -zein fraction was composed of a major 17 kDa protein band, while the -coixin fraction consisted of a mixture of - and -coixins.Polyclonal antibodies raised against C1 recognized C1 and C2 and cross-reacted strongly with the 22 kDa -zein, as did C4 and C5 antisera. The antiserum against -coixin showed strong cross-reaction with -zein. The homology between coixins and zeins was further investigated by using Southern hybridization analyses. The genomic DNA of maize and Coix were digested with several restriction enzymes and probed with cDNA clones representing 19 and 22 kDa -zeins as well as the 28 and 16 kDa -zeins. The Coix genome showed complex cross-hybridization sequences with the 22 kDa -zein cDNA, while no cross-hybridization was observed with the 19 kDa cDNA clone. The cDNA clone representing the 28 kDa -zein cross-hybridized with only one band of Coix genomic DNA, in contrast to the three bands observed in maize. This same Coix sequence also cross-hybridized with the cDNA clone representing the 16 kDa -zein. The relevance of these findings are discussed in the context of the origin of zein and coixin genes.     

Analysis of the effect of rye chromosomes 4R, 6R and telomeric heterochromatin on 7R on homologous chromosome pairing in pentaploid hybrids (AABBR,AABBD)

Summary Meiotic pairing in Triticum turgidum cv. Ma (4x) with a mean chiasmata frequency of 27.16 per cell was compared with chiasmata frequencies in its hybrids with several triticale strains, Chinese Spring wheat and its addition lines for Imperial rye chromosomes 4R and 6R. In hybrids between Ma and x Triticosecale cv. Rosner the chiasmata frequency was marginally reduced by an average of 1.25%, by 8.8% in hybrids with x Triticosecale cv. DRIRA HH and by 6.7% with DRIRA EE (lacking 90% telomeric heterochromatin from chromosome arm 7RL). In pentaploid hybrids between Ma and T. aestivum cv. Chinese Spring the reduction was an average of 10.30%, while addition lines with rye chromosome 6R reduced chiasmata frequencies by an average of 7.4% and rye addition line for 4R showed the greatest depression in chiasmata frequency in hybrids by a 25.04% reduction. An interchange difference involving long chromosome segments was observed between Ma and Rosner.Contribution No. 819 Ottawa Research Station     

Ultrastructural localization of ‘anixiopsin’: a lectin of the fungus Anixiopsis stercoraria (Hansen) Hansen

In order to localize the anixiopsin, a lectin of the keratinolytic fungus Anixiopsis stercoraria, the authors used a monospecific antiserum prepared by immunization of rabbits with their own erythrocytes coated in vitro with anixiopsin. In light and scanning electron microscopies, lectinic sites were visualized by means of latex microspheres sensitized with anti-rabbit IgG antibodies. In transmission electron microscopy using the IgG fraction of the rabbit anti-anixiopsin immune sera and protein A-gold, anixiopsin seemed mainly present on the outermost cell wall layer of the ascospores, in a pseudomembraneous structure dense to electrons. Implications of these results on physical and biological properties of the lectin are discussed.     

Spatial and temporal variation in carbon isotope discrimination in prairie graminoids

We present the results of a 5-year examination of variation in the stable carbon isotope composition () of three C₃ graminoid species from a Sandhills prairie: Agropyron smithii, Carex heliophila and Stipa comata. Although consistent species-specific patterns for mean were seen, there was also significant and substantial among-year and within-season variation in . A smaller contribution to variation in came from topographic variation among sampling sites. Effects of species, year, season and topography contribute to variation in in an additive manner. An association between climate and exists that is consistent with previous work suggesting that reflects the interplay between photosynthetic gas exchange and plant water relations. Within the growing season, the time over which integrates plant response to the environment ranges from days to months.     

‘Hard’ data for matrix modelling of Laminaria digitata (Laminariales,Phaeophyta) populations

The objective of the study was to produce a size-based matrix model of a Laminaria digitata (L.) Lamour. population. Hard data for insertion in the matrix were collected in a 9 year cohort analysis of size and age specific survival and fertility for a stand in south west Nova Scotia, Canada. The product of the square matrix containing these values and a column vector containing the densities of size classes was used to project the size class structure one year later. The projected estimates were found to fit empirical estimates with some confidence. In contrast, an age-based fertility life table wrongly predicted a population declining in density by 45% per year. The study supports, in theory, the use of size-based matrix models for management of harvested stands. In reality, the amount of work required to obtain hard data and the site specific nature of the projections may preclude the use of such an approach to broad scale management.     

Discovery,localization, and sequence characterization of molecular markers for the crown rust resistance genes <Emphasis Type="Italic">Pc38, Pc39</Emphasis>, and <Emphasis Type="Italic">Pc48</Emphasis> in cultivated oat (<Emphasis Type="Italic">Avena sativa</Emphasis> L.)

Molecular markers for the crown rust resistance genes Pc38, Pc39, and Pc48 in cultivated oat (Avena sativa L.) were identified using near-isogenic lines and bulked segregant analysis. Six markers for Pc48, the closest being 6 cM away, were found in a Pendek-39 × Pendek-48 (Pendek3948) population, but none was found in a Pendek-48 × Pendek-38 (Pendek4838) population. Three markers for Pc39 were found in the Pendek3948 population, one of which cosegregated with the gene. This same marker was found to be 6 cM away from the gene in an OT328 × Dumont (OT328Du) population. Nine markers for Pc38 were found in the Pendek4838 population, eight of which are within 2 cM of the gene. One other marker for Pc38 was found in the OT328Du population; however, comparative mapping suggests that the Pc38 region in OT328Du is in a different location than that in Pendek4838. A number of markers unlinked to the genes under study formed linkage groups in both the Pendek3948 and Pendek4838 populations. Four of these show homology or homoeology to each other and to the Pc39 region in Pendek3948. Two RFLP clones closely linked to Pc38 code for a putative leucine-rich repeat transmembrane protein kinase and a cre3 resistance gene analogue. This study provides information to support molecular breeding in oat, and contributes to ongoing research into genomic regions associated with fungal pathogen resistance.     

Expression of the β-subunit of β-conglycinin in seeds of transgenic plants

Soybean (Glycine max (L.) Merr.) seeds contain the storage protein -conglycinin, encoded by a multigene family. -Conglycinin consists of three subunits; , , and . A genomic clone for a -subunit of -conglycinin has been characterized by restriction-enzyme mapping and hybrid selected in-vitro translation followed by immunoprecipitation. In order to determine the developmental regulation of this -subunit gene, its expression was studied in seeds of transgenic petunia (Petunia hybrida) and tobacco (Nicotiana tabacum L.) plants. The -subunit expressed in seeds of petunia and tobacco was recognized by anti--conglycinin serum at a relative molecular mass of 53 000, equivalent to that of the native protein. Separation of the petunia-seed proteins by isoelectric focusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis showed that multiple isoelectric forms of the -subunit were produced. There was approximately a twofold variation in the accumulation of the -subunit protein in the mature seeds of transgenic petunia plants, each containing a single -subunit gene. However, the level of protein accumulation in mature seeds and the amount of -subunit mRNA in developing seeds was not correlated. Accumulation of the -subunit protein in transgenic seeds was less than the -subunit protein that accumulated in transgenic petunia seeds containing a single -subunit gene and less than the amount of the -subunit in mature soybean seeds which contain 8–13 -subunit genes. In transgenic tobacco plants, the accumulation of the -subunit protein in seeds was generally well correlated with the number of genes that were incorporated in the different transformants.Abbreviations kb kilobase - kDa kilodalton - M_r relative molecular mass - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis     

Correlations between in vivo resistance to Fusarium and in vitro response to fungal elicitors and toxic substances in carnation

Summary With the aim of ascertaining the existance of a correlation between in vivo resistance to Fusarium oxysporum f. sp. dianthi and in vitro response to fungal elicitors and toxic substances, phenylalanine ammonialyase and phytoalexin accumulation, on one hand, and resistance to culture filtrate, on the other, were assayed in in vitro cultures of three susceptible and four resistant Dianthus caryophyllus cultivars. Cultivars showing varying degrees of resistance in vivo either tolerated higher culture filtrate concentrations (Niki) or showed high PAL activity and phytoalexin production when treated with Fusarium elicitor (Duca), or responded positively to both treatments (Mei-Ling, Pulcino). No such responses were shown in tissue cultures of susceptible cultivars. The differential response to the fungal elicitor seemed to be highly specific as genetic differences between cultivars were not observed in tissue cultures treated with other biotic (Phytophthora infestans) and abiotic (HgCl₂) elicitors.Abbreviations FuCWC cell wall components from Fusarium oxysporum f. sp. dianthi race 2 - PhCWC cell wall components from Phytophthora infestans - PAL phenylalanine ammonia-lyase (EC 4.3.1.5)