Dirce Maria Carraro

Structural features and transcript-editing analysis of sugarcane (Saccharum officinarum L.) chloroplast genome

The complete nucleotide sequence of the chloroplast genome of sugarcane (Saccharum officinarum) was determined. It consists of 141,182 base-pairs (bp), containing a pair of inverted repeat regions (IRA, IRB) of 22,794xa0bp each. The IRA and IRB sequences separate a small single copy region (12,546xa0bp) and a large single copy (83,048xa0bp) region. The gene content and relative arrangement of the 116 identified genes (82 peptide-encoding genes, four ribosomal RNA genes, 30xa0tRNA genes), with the 16 ycf genes, are highly similar to maize. Editing events, defined as C-to-U transitions in the mRNA sequences, were comparable with those observed in maize, rice and wheat. The conservation of gene organization and mRNA editing suggests a common ancestor for the sugarcane and maize plastomes. These data provide the basis for functional analysis of plastid genes and plastid metabolism within the Poaceae. The sugarcane chloroplast DNA sequence is available at GenBank under accession NC005878.

Expression of putative pathogenicity-related genes in Xylella fastidiosa grown at low and high cell density conditions in vitro

Xylella fastidiosa is the causal agent of economically important plant diseases, including citrus variegated chlorosis and Pierces disease. Hitherto, there has been no information on the molecular mechanisms controlling X. fastidiosa-plant interactions. To determine whether predicted open reading frames (ORFs) encoding putative pathogenicity-related factors were expressed by X. fastidiosa 9a5c cells grown at low (LCD) and high cell density (HCD) conditions in liquid modified PW medium, reverse Northern blot hybridization and reverse transcription-polymerase chain reaction (RT-PCR) experiments were performed. Our results indicated that ORFs XF2344, XF2369, XF1851 and XF0125, encoding putative Fur, GumC, a serine-protease and RsmA, respectively, were significantly suppressed at HCD conditions. In contrast, ORF XF1115, encoding putative RpfF, was significantly induced at HCD conditions. Expressions of ORFs XF2367, XF2362 and XF0290, encoding putative GumD, GumJ and RpfA, respectively, were detected only at HCD conditions, whereas expression of ORF XF0287, encoding putative RpfB was detected only at LCD conditions. Bioassays with an Agrobacterium traG::lacZ reporter system indicated that X. fastidiosa does not synthesize N-acyl-homoserine lactones, whereas bioassays with a diffusible signal factor (DSF)-responsive Xanthomonas campestris pv. campestris mutant indicate that X. fastidiosa synthesizes a molecule similar to DSF in modified PW medium. Our data also suggest that the synthesis of the DSF-like molecule and fastidian gum by X. fastidiosa is affected by cell density in vitro.

From Tissue Samples to Tumor Markers

Changes in the general transcription profile have been observed through silencing and activating at the transcriptional level of genes in tumor cells. After the genomic era, molecular biology has changed, and new technologies have allowed assessing transcriptional alterations among different tissue types in a high-throughput manner. Microarray technology is one of the technologies that has contributed to improving our understanding about the defective molecular processes in cancer cells. In this chapter, we report issues about the selection of cDNA clones spotted on the platform, the purity of the tumor samples used in the microarray experiments, and the integrated database with important clinical information of patients that can be associated to a specific molecular portrait. Finally, we focus on the validation of candidate genes selected from microarray experiments through real-time RT-PCR and high-throughput tissue microarray analyses that dramatically facilitate testing of the potential molecular markers.

Gene expression profile of stromal cells from potential metastatic sites in breast cancer patients.

Methods Fibroblasts primary culture was established from 11 breast cancer patients. Expression analysis was evaluated in PT (n=4), N+ (n=3) and BM (n=4) through a customized cDNA microarray platform (4,800 ORESTES) analyzed by SAM (TMEV; FDR 0%) and functional analysis was performed using DAVID. Technical validation was performed in 6 samples and biological validation was performed in fibroblasts obtained from others 25 patients as evaluated by RT-qPCR.

Genome-wide profiling of copy number alterations in triple-negative breast cancer identifies a region at 19p13 associated with lymph node metastasis

Background The acquisition of somatic alterations (point mutations/ chromosomal rearrangements) underlies the hallmarks of cancer, generating genetic diversity that drives tumorigenesis. Advances in the study of cancer genomes revealed in solid tumors a complex pattern of copy number alterations (CNA), structural rearrangements, and aneuplodies. Breast cancer (BC) is the most common malignancy in females, being the leading cause of death by cancer. This heterogeneous disease is not fully understood yet; however, genomic studies have identified unique CNA patterns in different BC subtypes. Regarding the subtype triple-negative (TN; estrogen and progesterone receptors, and HER2 negative expression levels), only limited data are available on which genes or chromosome regions are involved in its initiation and progression.