Liziane Maria de Lima

Brazilian coffee genome project: an EST-based genomic resource

Coffee is one of the most valuable agricultural commodities and ranks second on international trade exchanges. The genus Coffea belongs to the Rubiaceae family which includes other important plants. The genus contains about 100 species but commercial production is based only on two species, Coffea arabica and Coffea canephora that represent about 70 % and 30 % of the total coffee market, respectively. The Brazilian Coffee Genome Project was designed with the objective of making modern genomics resources available to the coffee scientific community, working on different aspects of the coffee production chain. We have single-pass sequenced a total of 214,964 randomly picked clones from 37 cDNA libraries of C. arabica, C. canephora and C. racemosa, representing specific stages of cells and plant development that after trimming resulted in 130,792, 12,381 and 10,566 sequences for each species, respectively. The ESTs clustered into 17,982 clusters and 32,155 singletons. Blast analysis of these sequences revealed that 22 % had no significant matches to sequences in the National Center for Biotechnology Information database (of known or unknown function). The generated coffee EST database resulted in the identification of close to 33,000 different unigenes. Annotated sequencing results have been stored in an online database at http://www.lge.ibi.unicamp.br/cafe. Resources developed in this project provide genetic and genomic tools that may hold the key to the sustainability, competitiveness and future viability of the coffee industry in local and international markets.

Allelopathy, an alternative tool to improve cropping systems. A review

Known since ancient times, the phenomenon of allelopathy has recently received greater attention from researchers and farmers worldwide. Crop failures and low yields caused by the reseeding, overseeding, crop rotation and replanting of fruit trees in orchards are believed to be caused by allelopathic activity. Moreover, the expensive and environmentally impacting herbicides for weed control have motivated studies directed at developing cheaper, environmentally-friendly alternatives. Despite the difficulties in separating chemical interference from other mechanisms, more modern approaches have produced consistent and conclusive data on this topic. The release and dynamics of allelochemicals in the soil are discussed herein. Examples of allelopathic crops, the allelochemicals produced and their uses in cropping systems are also presented along with current research trends regarding allelopathy.

Plant Promoters: An Approach of Structure and Function

With current advances in genomics, several technological processes have been generated, resulting in improvement in different segments of molecular research involving prokaryotic and eukaryotic systems. A widely used contribution is the identification of new genes and their functions, which has led to the elucidation of several issues concerning cell regulation and interactions. For this, increase in the knowledge generated from the identification of promoters becomes considerably relevant, especially considering that to generate new technological processes, such as genetically modified organisms, the availability of promoters that regulate the expression of new genes is still limited. Considering that this issue is essential for biotechnologists, this paper presents an updated review of promoters, from their structure to expression, and focuses on the knowledge already available in eukaryotic systems. Information on current promoters and methodologies available for studying their expression are also reported.

Produtividade de grãos e óleo de genótipos de amendoim para o mercado oleoquímico

Top lines and cultivars of runner peanut genotypes were analyzed as to grain and oil yield, aiming further recommendation to edible or combustible oil market. The genotypes were cultivated during wet seasons for two years, at the city of Barbalha, state of Ceara, Brazil, by using a complete randomized block design with five replicates. The harvest took place from 110 to 135 days after planting. Pod, grain and oil yields were registered in each plot. Crude oil was extracted by soxhlet protocol, by using petroleum ether as a solvent. Afterwards, fatty acids were analyzed by gas chromatography. Based on yield tests, LViPE-06 and BRS Perola Branca showed the highest grain yields, with pod and seed yield averages of 3.04 kg ha-1 and 2.13 kg ha-1, respectively. These genotypes also stood out as to oil chemistry industry, based on oil content and ratio of oleic/ linoleic fatty acids, which were 51 and 1.9, respectively.

Variabilidade de progênies F2 de amendoim geradas por meio de seleção de genitores ISSR-divergentes

Marcadores ISSR foram utilizados para discriminar acessos intraespecificos de amendoim e utilizar os mais divergentes para geracao de variabilidade por meio de cruzamentos. Os acessos foram cultivados em casa de vegetacao e aos 20 dias apos o plantio foram coletadas folhas para extracao de DNA e posterior ensaios de PCR-ISSR. Dez primers foram utilizados gerando bandas mono e polimorficas. Dentre eles, os mais responsivos em termos de discriminacao dos acessos foram UBC-818 e UBC-842, que geraram, respectivamente, 14 e 12 bandas e taxa de polimorfismo de 64 e 83%. Os primers UBC-847 e UBC-858 geraram poucas bandas e mais de 60% foram monomorficas, de pouco valor para estudos de discriminacao de acessos por meio de marcadores do tipo ISSR. Tres grupos distintos foram formados no dendrograma, entre os quais, dois genotipos de maior divergencia, representados pela cultivar BR 1 (precoce, de porte ereto) e a linhagem LViPE-06 (tardio e rasteiro), foram selecionados para os trabalhos de hibridacao para gerar variabilidade genetica. As progenies F2 resultantes apresentaram larga variabilidade genetica, de grande contribuicao para os trabalhos de selecao no melhoramento do amendoim. Com base nos resultados obtidos, foi possivel confirmar a contribuicao das ferramentas moleculares como auxiliar nos trabalhos de selecao de amendoim.

Stable integration and expression of a cry1Ia gene conferring resistance to fall armyworm and boll weevil in cotton plants

BACKGROUND Boll weevil is a serious pest of cotton crop. Effective control involves applications of chemical insecticides, increasing the cost of production and environmental pollution. The current genetically modified Bt crops have allowed great benefits to farmers but show activity limited to lepidopteran pests. This work reports on procedures adopted for integration and expression of a cry transgene conferring resistance to boll weevil and fall armyworm by using molecular tools. RESULTS Four Brazilian cotton cultivars were microinjected with a minimal linear cassette generating 1248 putative lines. Complete gene integration was found in only one line (T0-34) containing one copy of cry1Ia detected by Southern blot. Protein was expressed in high concentration at 45 days after emergence (dae), decreasing by approximately 50% at 90 dae. Toxicity of the cry protein was demonstrated in feeding bioassays revealing 56.7% mortality to boll weevil fed buds and 88.1% mortality to fall armyworm fed leaves. A binding of cry1Ia antibody was found in the midgut of boll weevils fed on T0-34 buds in an immunodetection assay. CONCLUSION The gene introduced into plants confers resistance to boll weevil and fall armyworm. Transmission of the transgene occurred normally to T1 progeny. All plants showed phenotypically normal growth, with fertile flowers and abundant seeds.

Control of Sclerotium rolfsii in peanut by using Cymbopogon martinii essential oil

Essential oils of seven species were investigated in order to control peanut plants against white mold (Sclerotium rolfsii Sacc.). The assays were carried out by in vitro and in vivo assays. At first, fungitoxicity and suppression of oxalic acid diffusion (SOAD) bioassays were performed in order to evaluate the mycelial growth of fungus. Then, validation assays were carried out in greenhouse, involving inoculation of fungus in the seeds and further plant treatments with essential oil. Four isolates of S. rolfsii were tested in different oil concentrations. Cymbopogon martinii oil at 300 ppm inhibited the mycelia growth of S. rolfsii in 55% and also the number of sclerotia. In validation assay, we found that a single dose of C. martinii oil at 400 ppm reduced the rate of disease in 55%, confirming the in vitro assays. The follows traits: number of pods/plant, pod weight and harvest index increased in 57, 54, and 40%, respectively, in all C. martini oil treatments. These results demonstrate that C. martinii oil at low concentration may serve for new formulations in the treatment and prevention of white mold. Key words: Arachis hypogaea, disease control, toxicity, white mold.

Validating a probe from GhSERK1 gene for selection of cotton genotypes with somatic embryogenic capacity

Substantial progress is being reported in the techniques for plant transformation, but successful regeneration of some genotypes remains a challenging step in the attempts to transform some recalcitrant species. GhSERK1 gene is involved on embryo formation, and its overexpression enhances the embryogenic competence. In this study we validate a short GhSERK1 probe in order to identify embryogenic cotton genotypes using RT-qPCR and blotting assays. Cotton genotypes with contrasting somatic embryogenic capacity were tested using in vitro procedures. High expression of transcripts was found in embryogenic genotypes, and the results were confirmed by the RT-PCR-blotting using a non-radioactive probe. The regeneration ability was confirmed in embryogenic genotypes. We confirmed that GhSERK1 can be used as marker for estimating the somatic embryogenesis ability of cotton plants.

VARIABILIDADE GENOTÍPICA DE AMENDOIM EM RESPOSTA A ESTRESSE HÍDRICO BASEADA EM DESCRITORES BIOQUÍMICOS

Seven biochemical descriptors were used to estimate the genotypic variability of peanut in response to moderate water stress. Six genotypes, constituted by four lines and two cultivars, were grown in pots, each containing two plants. At 15 days after emergence (DAE), the treatment differentiation was carried out: Control-plants maintained with daily watering, and Stress-plants submitted to water stress by complete suspension of watering for 15 days. The experimental design was completely randomized with factorial scheme 6 x 2 (genotype x water treatments), with five replications. The biochemical variables evaluated were: catalase (CAT), ascorbate peroxidase (APX), guaiacol peroxidase (GPX), free proline, total carbohydrates, soluble proteins, and amino acids. Results obtained by biochemical analysis and estimation of genotypic variability indicated that proline is the most appropriate descriptor for selecting genotypes tolerant to water stress, which led to identification of L81V and L108V as promising lines for drought tolerance breeding program.

Bioactivity of basil ( Ocimum basicilum L.) on control of the spider mite ( Tetranychus urticae Koch.) in peanut

Essential plant oils contain biopesticides that could be used to control many crop pests. Tetranychus spp. are mites that cause damage to several crops and are primarily controlled by synthetic pesticides. Literature showed that mites can be controlled with essential oils of plants containing eugenol. In this work, we evaluated the bioactivity of basil (Ocimum basicilum) accessions for peanut-spider mites control based on molecular, biochemical and agronomic assays. RNA from four basil accessions, previously chosen by divergence genetic analysis, were used to estimate the expression of eugenol synthase (EGS I) transcripts, by semiquantitative and polymerase chain reaction (qPCR) assays. Chromatography was, thereafter, performed in order to estimate the eugenol concentration. Feeding bioassays were performed using basil leaf extracts in order to estimate oviposition and mortality of spider mites females. Finally, a validation assay was carried out in greenhouse, using peanut plants previously infested with spider mites and weekly sprayed with basil water-extract. One basil accession, OVRS, revealed high phytotoxicity to spider mite females, at 15% water-extract. The mortality rate was 75% and complete inhibition of fecundity was found in BOD assays. In the greenhouse assay, the most severe damage due to mite infestations was found to plant height, number of pods and pod yield, which were reduced to 28, 53 and 52% in non-treated plants (control). Considering that basil is a short-cycle plant, with easy reproduction and management, these results represent an accessible alternative to organic control spider mites in peanut.