One step shoot tip multiplication and rooting of Digitalis thapsi L.

Shoot tips from seedlings of Digitalis thapsi L. were cultured on Murashige and Skoog's medium and the effect of various auxins (2,4-D, NAA and IAA) were analyzed alone or in combination with cytokinis (BA and kinetin). Shoot multiplication and direct rooting of the new shoots were obtained after four weeks of culture in MS medium without hormones, but callus formation and the appearance of abnormal phenotypes were frequent. The addition of auxins to the cultures prevented the formation of callus but not the appearance of variant phenotypes. Both drawbacks could be avoided by combination of NAA or IAA with BA or kinetin. The best results for shoot multiplication and direct rooting were obtained with 0.5 mg l^-1 NAA and 0.1 or 0.5 mg l^-1 kinetin.Abbreviations BA 6-benciladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indole-3-acetic acid - Kin kinetin - NAA naphtalene acetic acid - MS Murashige and Skoog     

Enhanced immunocytochemical staining sensitivity in formalin-fixed and paraffin-embedded material by simultaneous use of PAP and ABC

The simultaneous use of two immunoperoxidase (IP) methods; e.g. the PAP (Peroxidase-antiperoxidase) and ABC (Avidin-Biotin) to localize a single antigen enhances the sensitivity of polyclonal antibodies (FVIII/RAg and laminin) or monoclonal antibodies (MABs) (against human B-, T-cells and macrophages) in routinely formalin fixed and in paraffin embedded material. The increased sensitivity was not accompanied by loss of specificity. With antibodies against Factor VIII related antigen (FVIII/RAg) and laminin the increased staining intensity of blood vessels were demonstrated in experimental rat transplantation tumors. The similar enhancement of immunocytochemical reactions was observed with biopsies of human brain tumors, where the monoclonal antibodies against white blood cells were used. The staining results were of comparable intensity as in snapfrozen tissue or after special embedding procedure for immunocytochemical purposes acc. to Bolton and Mesnard formol sucrose gum, sucrose paraffin; FSGSP).     

Genetic variation of recent Alu insertions in human populations

The Alu family of intersperesed repeats is comprised of ovr 500,000 members which may be divided into discrete subfamilies based upon mutations held in common between members. Distinct subfamilies of Alu sequences have amplified within the human genome in recent evolutionary history. Several individual Alu family members have amplified so recently in human evolution that they are variable as to presence and absence at specific loci within different human populations. Here, we report on the distribution of six polymorphic Alu insetions in a survey of 563 individuals from 14 human population groups across several continents. Our results indicate that these polymorphic Alu insertions probably have an African origin and that there is a much smaller amount of genetic variation between European populations than that found between other populations groups. Present address: Department of Pathology, Stanley S. Scott Cancer Center, Louisiana State University Medical Center, 1901 Perdido St., New Orleans, LA 70112 Correspondence to: M.A. Batzer     

Cloning and expression of the Neisseria meningitidis 5C outer membrane protein

The 5C outer membrane protein, one of the N. meningitidis class 5 proteins, was preferably expressed in bacteria isolated from the nasopharynx and its role in adhering to the mucosal cells and invading them as well as the development of anti-5C antibodies in healthy carriers was demonstrated. Anti-5C monoclonal antibodies are bactericidal in the presence of the human complement. The immunodominant region of the 5C protein is highly conserved among the different strains of N. meningitidis, and the opc gene, which encodes the protein, does not seem to show antigenic variations. Here the isolation of the opc gene from the Cuban strain B:4:P1.15 by PCR (Polymerase Chain Reaction) is presented. Under the regulation of the tryptophan promoter, the gene was cloned and sequenced in E. coli with a high level of expression and fused to the amino-terminal end of the interleukin-2 gene. In the dot-blot experiments, the presence of the gene in those strains which did not express the protein in the whole cell ELISA was also detectable.     

The age, growth and reproduction of Chondrostoma polylepis willkommi in a seasonal stream in the Guadalquivir River basin (southern Spain)

The age, growth and reproduction of Chondrostoma polylepis willkommi , an endemic cyprinid from the Iberian Peninsula, was studied for 2 years in a seasonal stream in the Guadalquivir River basin. Annual growth was low. Maximum ages observed were 5+ in males and 7+ in females. There was no significant difference in growth between sexes. Seasonal growth period started in May and continued for 2 to 6 months depending on age. The mean lengths of 1 + and 2+ age-groups decreased once the growth period had finished, probably related to reproductive stress and adverse ecological conditions, and this could explain the occurrence of Rosa Lee's phenomenon. Both sexes matured at the end of their second year of life (1+). There was a significant difference from 1: 1 in the overall sex ratio of 334 males to 464 females. Reproductive period started in March and lasted until May. Ch. p. willkommi was a multiple spawner that released two batches of eggs per female each year. The regression between fecundity ( F ) and fork length (l., mm) was: F =6.20 10³ L.^2.78. In females from age 2 + onwards, relative annual investment in somatic growth and reproduction was equivalent, implying that the same energy was allocated to reproduction as to maintenance/growth. Compared with other Ch. polylepis populations, the life-history patterns of this population, located in a small and seasonal stream in a southern latitude, were characterized by a low annual growth, a trade-off between reproduction and growth/maintenance, early maturity, low number of age-groups and high fecundity from multiple spawnings.     

Selection of anthocyanin-accumulating potato (Solanum tuberosum L.) cell lines from calli derived from seedlings produced by gamma-irradiated seeds

Callus cell lines of potato (Solanum tuberosum L. cv. Zarevo) were obtained from seedlings germinated from gamma-irradiated seeds (200 Gy). Some of these cell lines produce red-violet pigments which were identified as acylated anthocyanins. The major anthocyanin was determined to be peonidin 3-O-[6-O-(4-O-E-p-coumaroyl-rhamnosyl)-glucoside]-5-O-glucoside (peonanin). Single cell-derived protoclones from non-pigmented protoplasts sometimes also gave rise to pigmented cell clusters thus indicating that the changes in the expression of the anthocyanin pathway can also occur after the stage of initial callus induction.     

Deep-water macroalgae from the Canary Islands: new records and biogeographical relationships

Due to the geographical location and paleobiogeography of the Canary Islands, the seaweed flora contains macroalgae with different distributional patterns. In this contribution, the biogeographical relations of several new records of deep-water macroalgae recently collected around the Canarian archipelago are discussed. These areBryopsidella neglecta (Berthold) Rietema,Discosporangium mesarthrocarpum (Meneghini) Hauck,Hincksia onslowensis (Amsler et Kapraun) P. C. Silva,Syringoderma floridana Henry,Peyssonnelia harveyana J. Agardh,Cryptonemia seminervis (C. Agardh) J. Agardh,Botryocladia wynnei Ballantine,Gloiocladia blomquistii (Searles) R. E. Norris,Halichrysis peltata (W. R. Taylor) P. Huvé et H. Huvé,Leptofauchea brasiliensis Joly, andSarcodiotheca divaricata W. R. Taylor. These new records, especially those in the Florideophyceae, support the strong affinity of the Canary Islands seaweed flora with the warm-temperate Mediterranean-Atlantic region. Some species are recorded for the first time from the east coast of the Atlantic Ocean, enhancing the biogeographic relations of the Canarian marine flora with that of the western Atlantic regions.     

Selection of synthetic proteins to modulate the human frataxin function

Frataxin is a kinetic activator of the mitochondrial supercomplex for iron-sulfur cluster assembly. Low frataxin expression or a decrease in its functionality results in Friedreich's Ataxia (FRDA). With the aim of creating new molecular tools to study this metabolic pathway, and ultimately, to explore new therapeutic strategies, we have investigated the possibility of obtaining small proteins exhibiting a high affinity for frataxin. In this study, we applied the ribosome display approach, using human frataxin as the target. We focused on Affi_224, one of the proteins that we were able to select after five rounds of selection. We have studied the interaction between both proteins and discussed some applications of this specific molecular tutor, concerning the modulation of the supercomplex activity. Affi_224 and frataxin showed a K_D value in the nanomolar range, as judged by surface plasmon resonance analysis. Most likely, it binds to the frataxin acidic ridge, as suggested by the analysis of chemical shift perturbations (nuclear magnetic resonance) and computational simulations. Affi_224 was able to increase Cys NFS1 desulfurase activation exerted by the FRDA frataxin variant G130V. Importantly, Affi_224 interacts with frataxin in a human cellular model. Our results suggest quaternary addition may be a new tool to modulate frataxin function in vivo. Nevertheless, more functional experiments under physiological conditions should be carried out to evaluate Affi_224 effectiveness in FRDA cell models.     

Age structure, growth and reproduction of Leuciscus pyrenaicus in an intermittent stream in the Guadalquivir river basin, southern Spain

The age, growth and reproduction of Leuciscus pyrenaicus (Günther, 1868), an endemic cyprinid from the Iberian Peninsula, was studied from November 1987 to September 1989 in a small seasonal tributary of the Guadalquivir river basin. Maximum fork lengths observed were a 160 mm male with six scale annuli and a 171 mm female aged 7 +. Maximum ages observed were 7 + in males and 8 + in females. There were no significant differences in the annual growth increments between sexes. Seasonal growth period started in March and continued for 5 to 6 months. Mean lengths of 1 + specimens onwards diminished during summer and/or autumn. Males and females matured in their third and fourth year of life respectively. The overall sex ratio (272 males: 310 females) differed significantly from equality. Spawning began in May and ended in July. L. pyrenaicus is a multiple spawner that releases a minimum of two batches of eggs per female each year. Eggs in each batch were similar in both size (egg diameter) and number released. The relationship between fecundity (Fee) and fork length (mm) was represented by the formula: Fec=1.96 10⁻³ L^2.50.