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† Background and Aims The Mediterranean region is of prime importance to biodiversity at a global level, mainly due to the abundance of endemic plant species. However, information about these species is still scarce, especially at the genetic level. In this paper the first assessment is reported of the genetic structure of Centaurea horrida (Asteraceae), an endemic, sea-cliff-dwelling plant from Sardinia. † Methods The study was conducted on seven populations covering the entire natural range of the species by means of SSR (microsatellite) markers. † Key Results A considerable amount of genetic variation was found (average He ¼ 0.603‐0.854), together with a medium-high differentiation among populations, as estimated both by FST (0.123) and RST (0.158). Both Bayesian analysis and AMOVAwere employed to detect genetic structuring in this species. The results suggest that the origins of the current populations of C. horrida lie in two gene pools. † Conclusions Despite the restricted range, C. horrida displays high levels of genetic diversity, structured in such a way that three management units could be deemed viable for its conservation. The protected status of the species will probably suffice to prevent the impoverishment of its genetic resources.

Arabidopsis thaliana seedlings as measured by an electrolyte leakage assay, have been found to be extremely sensitive to high temperature stress as compared to a high temperature tolerant variety (Tracy) of soybean. Over 50% ion leakage occurred in Arabidopsis leaves during a 15-minute exposure to 50°C, indicating a heat killing time of less than 15 minutes. In contrast, the heat killing time for soybean at 50°C was over five times longer. When soybean or Arabidopsis seedlings in culture plates were exposed to 37°C for 2 hours and then returned to 23°C, they suffered no apparent short-term or long-term damage. Soybean seedlings given a 42°C, treatment for 2 hours also showed no damage. Arabidopsis seedlings after a 42°C treatment for 2 hours showed no apparent immediate damage, but 48 hours after return to 23°C severe damage symptoms were visible and after 96 hours all the seedlings were dead. Both soybean and Arabidopsis seedlings synthesize heat shock proteins (hsps) when exposed to 42°C for 2 hours. The hsps synthesized are of similar molecular weights, although the relative abundances of the different size classes are very different in the two plants. Even though hsps are produced in Arabidopsis seedlings after a 2 hour exposure to 42°C their presence is not sufficient for the seedlings to recover from the effects of rhe heat shock when returned to 23°C. Our results show that Arabidopsis has a heat sensitive genotype. This along with its other characteristics should make it a good model system in which to assay in transgenic plants, the functions of homologous and heterologous genes that might be candidates for determining heat tolerance in plants.

Fish have the ability to compensate for set-backs in growth as a result of fasting. When food levels are restored, growth in these fish can increase over and above normal rates. This phenomenon, known as "compensatory growth", has been studied with respect to enhancing food conversion efficiency. However, the mechanisms by which food intake activates an increase in somatic growth, and especially in muscle growth, are not well understood. In this study, we report first on the isolation of two complete cDNAs sequences encoding sea bass (Dicentrarchus labrax) myostatin and fibroblast growth factor 6 (FGF6), which have been shown to be major genetic determinants of skeletal muscle growth. The open reading frames of myostatin (376 amino acids) and FGF6 (209 amino acids) showed 97-63% and 87-62% sequence identity with other vertebrate myostatins and FGF6s, respectively. We also report on the expression profile of myostatin and FGF6 in sea bass skeletal muscle in response to different feeding regimens, as quantified by real-time RT-PCR. Nutritional status significantly influenced the myostatin expression levels in muscle, inducing an up-regulation during fasting and a down-regulation during the recovery from fasting, whereas the muscular FGF6 mRNA levels were not significantly affected by the feeding status of the animals. These findings suggest that myostatin has an inhibitory role in muscle growth in response to different feeding regimens, whereas FGF6 is not involved in the muscle compensatory growth induced by refeeding.

In this report we present the characterization of ovarian neoplasms including benign tumors, borderline tumors, and invasive carcinomas in order to assess whether a sharing of cytogenetic abnormalities is present in all three types of tumors. A cohort of 114 newly diagnosed and untreated ovarian epithelial tumors were analyzed by cytogenetic and molecular cytogenetic approaches with probes specific for chromosome 6. Three groups of chromosome abnormalities were identified: the first group included abnormalities common to all tumor classes (losses of chromosomes 6, 8, 10, 11, 15, 16, 17, 18, 19, 20, 21, 22, and X; gains of chromosomes 1, 3, 5, and 12; 6q24 approximately qter deletions); the second group presented specific abnormalities present in malignant but not in benign tumors (losses of chromosomes 2, 7, 13, and 14; gains of chromosome 4 and chromosome markers); and the last group included abnormalities unique to invasive carcinomas (loss of chromosome 4; gains of chromosomes 2, 7, 8, 9, 10, 16, 17, 18, 19, 20, and 21; 6q16 approximately q24 deletions; rearrangements of 3p, 3q, 13q, and 21q regions). The presence of shared chromosomal alterations in all three types of ovarian neoplasms investigated in this report seems therefore to suggest a progression model for these types of tumors.

We identified 15 patients with acute hepatitis C (AHC) among 29 healthy volunteers participating in 2 consecutive pharmacokinetics studies. Molecular techniques were used to determine the relatedness of viral strains, whereas clinical and virologic follow-up was started to establish the course and outcome of the acute infection. After presentation, serum liver enzymes and HCV RNA were monitored weekly for 4 months, then monthly for at least 12 months. Liver biopsy was performed 6 to 12 months after AHC diagnosis. Phylogenetic analysis of coding regions for the envelope glycoproteins E1 and E2 was performed. At presentation, all 15 patients tested HCV RNA-positive and had HCV genotype 2c. Phylogenetic analysis indicated a common source of infection. Fourteen patients agreed to be followed prospectively. Infection resolved spontaneously in 8 patients, HCV RNA becoming undetectable by 4 to 5 months after the presumed time of infection in 5 of them and by 8, 13, and 24 months in the remaining 3. Six patients developed chronic infection. Liver biopsies performed in 9 subjects who were HCV RNA-positive 6 months after AHC diagnosis revealed that the prevalent histologic finding was lobular inflammation. In conclusion, our homogeneous cohort showed a wide spectrum of clinical, virologic and histologic features, and, more importantly, short-term outcome differed noticeably despite the common source of infection.

The routes through which Norway spruce recolonized the Alps after the last ice age were investigated at the genetic level. Seven populations along the Alpine range plus one Apennine population were characterized for seven sequence-characterized amplified region (SCAR) loci, detecting an overall F-ST = 0.118. This rather high value for forest species reflects an uneven distribution of genetic variability, and was analysed through different statistical methods. Alternative hypotheses were tested under the isolation-by-distance model and using the analysis of molecular variance (amova) frame. We conclude that the hypothesis of the existence of a glacial refugium in the Apennines should be rejected, while a putative relict population is identified in the Maritime Alps. The Alpine range of Norway spruce appears to be split in two parts across a north-south line. The results are discussed in comparison with data based on morphological markers, isozymes, chloroplast microsatellites and mitochondrial markers.

Genetic factors controlling tolerance to the herbicide Alachlor in maize were localised by means of two different strategies. In the first approach, backcross (BC) plants, derived from pollen which had been subjected to selective pressure for resistance to the herbicide, were analysed for segregation distortion at 47 RFLP loci and compared to BC plants obtained from non-selected pollen. Preferential transmission of five chromosomal regions where putative QTLs (Quantitative Trait Loci) are localised was revealed in the BC plants from selected pollen. A second approach was based on a classical linkage analysis for segregation of the same set of RFLPs and factors controlling the trait, in a BC population of 210 individuals, by means of regression analysis. This study detected seven significant loci in four genomic regions. Overall, two loci revealed both segregation distortion and association with the expression of the trait, indicating linkage to genes expressed in both gametophytic and sporophytic phase. Three chromosomal regions appeared to carry factors involved in plant tolerance to Alachlor which are not expressed in pollen. Conversely, three loci were linked to factors selectable in pollen, but did not reveal significant association with tolerance in the plant in the segregating populations.

During a screen for mutants with defective germination, a new phenotype was observed consisting of red pigmentation of the embryonic axis in the dormant seed. Segregation ratios, as determined in F 2 and back-crossed progeny, indicate that the phenotype is due to a recessive single gene mutation that has been symbolized rea to denote red embryonic axis. A closer inspection of the rea phenotype revealed that the mutant is occasionally viviparous, indicating a defect in abscisic acid (ABA) metabolism. The mutation probably affects ABA sensitivity since no difference in ABA content was detected in mutant versus normal tissues. Moreover, when immature mutant and wild-type embryos were incubated on media containing 10 μM ABA, only the mutants germinated. ABA-regulated gene expression in rea embryos differed from that of embryos of the viviparous mutant vp1 which does not respond to the inhibitory action of ABA at the level of immature embryo germination. These results, therefore, indicate that the two genes exert a different role in the control of embryogenesis.

The dissection of the complex genetic program that regulates the male gametophytic phase can be accomplished by means of mutant analysis. The defective endosperm with gametophytic function (de-ga) mutants of maize are a category of monogenic viable gametophytic mutants in which the mutation also affects the development of the endosperm, causing a reduction of kernel size more or less severe depending on the mutant. As for the gametophytic effect of the mutation on microsporogenesis and/or on pollen tube growth, it can be easily evaluated by means of simple statistical analysis of the distortion from Mendelian segregation ratios in heterozygous (de-ga/+) plants (Ottaviano et al., 1988).

Densely saturated genetic maps of neutral genetic markers are a prerequisite either for plant breeding programs to improve quantitative traits in crops or for evolutionary studies. cDNA and genomic clones from maize were utilized to initiate the construction of a RFLP linkage map in Sorghum bicolor. To this purpose, an F2 population was produced from starting parental lines IS 18729 (USA) and IS 24756 (Nigeria) that were differentiated with regard to many morphological and agronomical traits. A total of 159 maize clones were hybridized to the genomic DNA of the two parents in order to detect polymorphism: 154 probes hybridized to sorghum and 58 out of these were polymorphic. In almost all of the cases hybridization patterns were similar between maize and sorghum. The analysis of the segregation of 35 polymorphic clones in an F2 population of 149 individuals yielded five linkage groups. The three principal ones recall regions of maize chromosomes 1, 3 and 5: in general, colinearity was maintained. A possible inversion, involving a long region of maize chromosome 3, was detected. Simulations were also performed to empirically obtain a value for the lowest number of individuals of the F2 population needed to obtain the same linkage data.

The success of a plant breeding program largely depends on the genetic composition of the base population used to select improved varieties and on the use of efficient selection procedures. The conventional approach offers a large spectrum of methods allowing increase of genetic variability and efficiency of selection. Practical results are well documented by the remarkable genetic gains obtained for many crops (Russell, 1974; Duvick, 1981; Borlaug, 1983). However, recent advances in biotechnology offer the possibility of increasing the efficiency of the conventional selection procedures and of enriching the genetic pool of a crop species by adding genes from alien species or obtained from in vitro manipulation. In this context, pollen technology can play an important role.

In contrast to sporophytic tissues, mature pollen of higher plants does not synthesize the typical set of heat-shock proteins (HSPs) in response to a marked temperature upshift. Immature grains, however, seem able to do so, at least partially. We investigated the characteristics of HSP synthesis throughout the male gametophytic phase in maize and compared gametophytic and sporophytic heat-shock responses. One-dimensional Sodium dodecyl sulfate-polyacryl-amide gel electrophoresis technique (SDS-PAGE) of newly synthesized proteins revealed that immature pollen synthesizes HSPs, some of which are not induced in sporophytic tissues. The heat-shock response appeared to be related to microgametophytic developmental stages. The strongest response was found in uninucleate microspores: at this stage, in addition to the sporophytic 102, 84, 72, and 18 kD HSPs, three other polypeptides of 74, 56, and 46 kD were observed. In the binucleate and trinucleate stages, only a reduced synthesis of few HSPs could be induced, and differences between genotypes were observed. In germinating pollen, HSP synthesis was not induced under a voriety of heat-stress conditions; however, the consti-tutive synthesis of two polypeptides of the same molecular weight, 72 and 64 kD, as two HSPs was observed. The biological significance of these results is discussed.

Gametophytic selection for important sporophytic traits such as resistence to environmental stresses and to pathogenic toxins has been proposed as a very efficient tool for the improvement of plant breeding methods1,2,3, on the grounds of the existence of an extensive genetic overlap between the gametophytic and sporophytic phases4,5,6.

To determine the extent of gametophytic gene expression and the type of transcription, haploid or haplo-diploid, of the genes, isozymes were used as genetic markers. Fifteen enzymatic systems, including thirty-four isozymes, were studied. The determination of the type of expression of genes coding for multimeric enzymes was based on the comparison of electrophoretic patterns of pollen and of sporophytic tissues from plants heterozygous for electrophoretic mobility: if gene expression in pollen is of a gametophytic (haploid) origin, pollen, unlike the sporophyte, would reveal only the parental homomultimeric bands. The enzymes analyzed can be grouped in three categories according to type of gene expression: i) enzymes present in both pollen and sporophyte, coded by the same gene with haplo-diploid expression; ii) enzymes controlling analogous functions in pollen and sporophyte, coded by different genes, expressed in only one of the two phases; iii) enzymes present in two or more forms in the sporophyte and only in one form in the gametophyte. The data allow the proportion of haplo-diploid gene expression in the loci examined to be estimated at 0.72; 0.22 and 0.06 being the proportions attributable to the sporophytic and gametophytic domains, respectively.

The basic prerequisite for an efficient breeding program to improve levels of resistance to pathogens in plants is the identification of genes controlling the resistance character. If the response to pathogens is under the control of a multilocus system, the utilization of molecular markers becomes essential. Stalk and ear rot caused by Gibberella zeae is a widespread disease of corn: resistance to G. zeae is quantitatively inherited. Our experimental approach to understanding the genetic basis of resistance to Gibberella is to estimate the genetic linkage between available molecular markers and the character, measured as the amount of diseased tissue 40 days after inoculation of a suspension of Fusarium graminearum, the conidial form of G. zeae, into the first stalk internode. Sensitive and resistant parental inbreds were crossed to obtain F1 and F2 populations: the analysis of the segregation of 95 RFLP (restriction fragment length polymorphism) clones and 10 RAPD (random amplified polymorphic DNA) markers was performed on a population of 150 F2 individuals. Analysis of resistance was performed on the F3 families obtained by selfing the F2 plants. Quantitative trait loci (QTL) detection was based either on analysis of regression coefficients between family mean value and allele values in the F2 population, or by means of interval mapping, using MAPMAKER-QTL. A linkage map of maize was obtained, in which four to five genomic regions are shown to carry factors involved in the resistance to G. zeae.