Agrobacterium-mediated transformation of the CrDAT gene and selection of transgenic periwinkle lines with a high vincristine accumulation

Catharanthus roseus contains vincristine and vinblastine, which are outstanding drugs for cancer. In the biosynthetic pathways of terpenoid indole alkaloids (TIAs) in C. roseus, deacetylvindoline 4-O-acetyltransferase (DAT) is a key enzyme that catalyses the last reaction of vindoline biosynthesis to form vinblastine and vincristine. In this study, the CrDAT transgene was transferred into the periwinkle by Agrobacterium-mediated transformation and generated transgenic periwinkle lines with an increase in vincristine accumulation. The C. roseus DAT gene was introduced into C. roseus plants and it was confirmed that CrDAT was successfully transferred into the genome of periwinkle plants and efficiently translated to synthesise recombinant DAT protein. Four transgenic periwinkle lines in T1 generation, T1-1, T1-3, T1-6, and T1-7, expressed recombinant DAT protein with the total protein content in the range of 2.86 μg.mg^?1 to 5.12 μg.mg^?1. Moreover, the vincristine contents of four transgenic lines increased by 1.63?2.48-fold compared to non-transgenic plants, ranging from 6.91 µg.g^?1 (fresh weight) to 10.53 µg.g^?1 (fresh weight). The T1-1 line had the highest vincristine content. Hence, the overexpression of the recombinant DAT protein can improve the vincristine accumulation of transgenic C. roseus plants.

Abbreviation: CrDAT - Catharanthus roseus Deacetylvindoline-4-O-Acetyl Transferase; D4H - Deacetoxyvindoline 4-hydroxylase; ELISA - Enzyme-Linked Immunosorbent Assay Monoterpene indole alkaloid; T0, T1 - Generations of transgenic plants; TIAs - Terpenoid indole alkaloids; WT- The wild-type tobacco plants (non transgenic plant); 35S - Cauliflower mosaic virus 35S promoter     

The optimal nutrition of maize in the hungarian national long‐term field experimental network

In Hungary, maize is grown on 1 million ha and occupies more than 20% of the arable land. The rich assortment of maize cultivars of different vegetation periods and different responses to nutritional effects, water supply etc. gives the growers the possibility to choose the cultivars suiting best the site characteristics (Jolânkai et al. 1999). Among the cereals maize has the highest genetical potential. To utilize its yield and quality potential, soil types of high nutrient content and regular nutrient supply are required (Gyõrffy, 1979). Both over‐ and under‐fertilization have an unfavourable effect on the yield and quality of maize (Debreczeni, 1985). Crops can be supplied with the appropriate nutrient amounts only with the knowledge of soil characteristics in the different agro‐ecological regions (nutrient content, water supply, soil compactness, pH, nutrient supplying capacity etc.). In Hungary, a network of long‐term field fertilization trials with uniform fertilizer treatments has been maintained at nine experimental sites representing different agro‐ecological regions of the country. This experimental network gives a basis to test the nutrient responses of our main crops and calibrate their optimal nutrient supply (Kismányoky, 1991).     

Circumsporozoite protein of Plasmodium vivax: gene cloning and characterization of the immunodominant epitope

The gene encoding the circumsporozoite (CS) protein of the human malaria parasite Plasmodium vivax has been cloned. The deduced sequence of the protein consists of 373 amino acids with a central region of 19 tandem repeats of the nonapeptide Asp-Arg-Ala-Asp/Ala-Gly-Gln-Pro-Ala-Gly. A synthetic 18-amino acid peptide containing two tandem repeats binds to a monoclonal antibody directed to the CS protein of Plasmodium vivax and inhibits the interaction of this antibody with the native protein in sporozoite extracts. The portions of the CS gene that do not contain repeats are closely related to the corresponding regions of the CS genes of two simian malarias, Plasmodium cynomolgi and Plasmodium knowlesi. In contrast, the homology between the CS genes of Plasmodium vivax and Plasmodium falciparum, another malaria parasite of humans, is very limited.     

Expression in Escherichia coli of open reading frame gene segments of HTLV-III

Human T-cell lymphotropic virus type III (HTLV-III), the causative agent of the acquired immune deficiency syndrome (AIDS), was recently isolated and its genomic structure analyzed by DNA cloning methods. In the studies reported here a combined cloning and expression system was used to identify HTLV-III encoded peptides that react immunologically with antibodies in sera from AIDS patients. Cloned HTLV-III DNA was sheared into approximately 500-base-pair fragments and inserted into an "open reading frame" expression vector, pMR100. The inserted DNA was expressed in Escherichia coli transformants as a polypeptide fused to the lambda CI protein at its amino terminus and to beta-galactosidase at its carboxyl terminus. Sera from AIDS patients containing antibodies to HTLV-III were then used to screen for immunoreactive fusion proteins. Twenty clones, each specifying a fusion protein strongly reactive with AIDS serum, were identified. DNA sequence analysis indicated that the HTLV-III fragments were derived from the open reading frame DNA segments corresponding to the gag and pol gene coding regions and also the large open reading frame region (env-lor) located near the 3' end of the viral genome.     

Repeated physical disturbances and the stability of sub‐tropical coral communities in Hong Kong,China

• 1. This paper reports on the dynamics of the coral community structure at A Ma Wan (AMW) and A Ye Wan (AYW) in Tung Ping Chau, Hong Kong, focusing on data collected before and after the summer typhoon seasons in 1997 to 1999. This period (1999) experienced the highest frequency of severe tropical cyclones to hit Hong Kong with one cyclone being the strongest to hit in the last 23 years (1984 to 2006).
• 2. This is part of a long‐term monitoring programme of subtropical coral communities that has been set up in AMW since May 1997 and AYW since May 1998 with nine fixed position 40 m long transects and 0.5 m × 0.5 m permanent quadrats laid at 5 m intervals along each transect. A total of 45 scleractinian coral species from 21 genera (12 families) was recorded, with 40 species in AMW and 32 species in AYW recorded at the beginning of the study.
• 3. Multidimensional scaling ordinations showed distinct spatial variations in the coral communities within and between sites. These variations were probably a result of the differential effects of the repeated cyclone impacts on the communities. Cyphastrea serailia, Goniopora lobata, Montipora peltiformis and Pavona decussata were the dominant species most strongly affected by cyclone impacts at AMW, experiencing a 6.2–16.2% loss of mean area cover during the study period. Some dominant species (e.g. P. decussata) in AYW also experienced a 6.5–14.3% loss of mean area cover within the same period.
• 4. Overall, these coral community structures appeared relatively stable and resistant to repeated physical disturbances. However, they may have low resilience towards more severe disturbances in the future. Preserving the resilience of coral communities/species in Tung Ping Chau should be one of the main goals of coral conservation strategies for the island.

Copyright © 2008 John Wiley & Sons, Ltd.     

Long-term high-titer neutralizing activity induced by octameric synthetic HIV-1 antigen

A titer for homologous viral neutralization activity (greater than 1:19,683) was observed after a 3.5-year immunization period with an octameric, branching peptide representing the principal neutralizing determinant (PND) of the human immunodeficiency virus-1IIIB envelope protein. Booster immunizations elicited persistent and potent antibodies in guinea pigs, exceeding responses produced by a conventional bovine serum albumin conjugate by 100-fold. Peptide length, central presentation of a conserved sequence, and inclusion of an upstream sequence contributed to immunogenicity. Titers (greater than 1:1,000) of heterotypic neutralizing antibodies also developed. Octameric PND peptides are a promising approach for an acquired immunodeficiency syndrome (AIDS) vaccine.     

Effect of mineral and organic fertilization on soil organic carbon content as well as on grain production of cereals in the IOSDV (ILTE) long-term field experiment,Keszthely, Hungary

The study was conducted in the international mineral and organic nitrogen fertilization trial (IOSDV) located in Keszthely in the western part of Hungary. The soil class of the study site was Ramann-type brown forest soil (Eutric Cambisol). The factors of the experiment are the increasing rate of mineral N fertilization and the complementary application of different forms of organic fertilizers in a three-course crop rotation (maize, winter wheat and winter barley) set up in 1983. The organic carbon content of the soil (C_org%) definitely increased after application of organic manure (OM), similarly to the average yield of the cultivated crops. After application of OM the increase in C_org content was 0.20%, and after straw + complementary N application it was 0.12%, compared with plots without organic manures. Mineral N fertilizer did not significantly influence the C_org content in soil over this period. Depending on the type of crop, the specific year and the N content of the soil, the combined application of mineral fertilizers and organic manures resulted in a 12–17% increase in crop yield, in general, whereas at maximum yields the increase accounted for 5–10%.