RESUMO
Anthonomus grandis Boheman is a key pest in cotton crops in the New World. Its larval stage develops within the flower bud using it as food and as protection against its predators. This behavior limits the effectiveness of its control using conventional insecticide applications and biocontrol techniques. In spite of its importance, little is known about its genome sequence and, more important, its specific expression in key organs like the midgut. Total mRNA isolated from larval midguts was used for pyrosequencing. Sequence reads were assembled and annotated to generate a unigene data set. In total, 400,000 reads from A. grandis midgut with an average length of 237 bp were assembled and combined into 20,915 contigs. The assembled reads fell into 6,621 genes models. BlastX search using the NCBI-NR database showed that 3,006 unigenes had significant matches to known sequences. Gene Ontology (GO) mapping analysis evidenced that A. grandis is able to transcripts coding for proteins involved in catalytic processing of macromolecules that allows its adaptation to very different feeding source scenarios. Furthermore, transcripts encoding for proteins involved in detoxification mechanisms such as p450 genes, glutathione-S-transferase, and carboxylesterases are also expressed. This is the first report of a transcriptomic study in A. grandis and the largest set of sequence data reported for this species. These data are valuable resources to expand the knowledge of this insect group and could be used in the design of new control strategies based in molecular information.
Assuntos
Transcriptoma , Gorgulhos/genética , Sequência de Aminoácidos , Animais , Simulação por Computador , Sistema Digestório/metabolismo , Larva/genética , Larva/metabolismo , Dados de Sequência Molecular , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Gorgulhos/crescimento & desenvolvimento , Gorgulhos/metabolismoRESUMO
Oligonucleotide-based microarrays with accurate gene coverage represent a key strategy for transcriptional studies in orphan species such as sunflower, H. annuus L., which lacks full genome sequences. The goal of this study was the development and functional annotation of a comprehensive sunflower unigene collection and the design and validation of a custom sunflower oligonucleotide-based microarray. A large scale EST (>130,000 ESTs) curation, assembly and sequence annotation was performed using Blast2GO (www.blast2go.de). The EST assembly comprises 41,013 putative transcripts (12,924 contigs and 28,089 singletons). The resulting Sunflower Unigen Resource (SUR version 1.0) was used to design an oligonucleotide-based Agilent microarray for cultivated sunflower. This microarray includes a total of 42,326 features: 1,417 Agilent controls, 74 control probes for sunflower replicated 10 times (740 controls) and 40,169 different non-control probes. Microarray performance was validated using a model experiment examining the induction of senescence by water deficit. Pre-processing and differential expression analysis of Agilent microarrays was performed using the Bioconductor limma package. The analyses based on p-values calculated by eBayes (p<0.01) allowed the detection of 558 differentially expressed genes between water stress and control conditions; from these, ten genes were further validated by qPCR. Over-represented ontologies were identified using FatiScan in the Babelomics suite. This work generated a curated and trustable sunflower unigene collection, and a custom, validated sunflower oligonucleotide-based microarray using Agilent technology. Both the curated unigene collection and the validated oligonucleotide microarray provide key resources for sunflower genome analysis, transcriptional studies, and molecular breeding for crop improvement.
Assuntos
Helianthus/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Etiquetas de Sequências Expressas , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologiaRESUMO
Sequence analysis of the 16S ribosomal RNA gene was used to study bacterial diversity of a pristine forest soil and of two cultures of the same soil enriched with cellulolytic bacteria. Our analysis revealed high bacterial diversity in the native soil sample, evidencing at least 10 phyla, in which Actinobacteria, Proteobacteria and Acidobacteria accounted for more than 76% of all sequences. In both enriched samples, members of Proteobacteria were the most frequently represented. The majority of bacterial genera in both enriched samples were identified as Brevundimonas and Caulobacter, but members of Devosia, Sphingomonas, Variovorax, Acidovorax, Pseudomonas, Xanthomonas, Stenotrophomonas, Achromobacter and Delftia were also found. In addition, it was possible to identify cellulolytic taxa such as Acidothermus, Micromonospora, Streptomyces, Paenibacillus and Pseudomonas, which indicates that this ecosystem could be an attractive source for study of novel enzymes for cellulose degradation.