A novel linkage map of sugarcane with evidence for clustering of retrotransposon-based markers.

BACKGROUND: The development of sugarcane as a sustainable crop has unlimited applications. The crop is one of the most economically viable for renewable energy production, and CO2 balance. Linkage maps are valuable tools for understanding genetic and genomic organization, particularly in sugarcane due to its complex polyploid genome of multispecific origins. The overall objective of our study was to construct a novel sugarcane linkage map, compiling AFLP and EST-SSR markers, and to generate data on the distribution of markers anchored to sequences of scIvana_1, a complete sugarcane transposable element, and member of the Copia superfamily. RESULTS: The mapping population parents ('IAC66-6' and 'TUC71-7') contributed equally to polymorphisms, independent of marker type, and generated markers that were distributed into nearly the same number of co-segregation groups (or CGs). Bi-parentally inherited alleles provided the integration of 19 CGs. The marker number per CG ranged from two to 39. The total map length was 4,843.19 cM, with a marker density of 8.87 cM. Markers were assembled into 92 CGs that ranged in length from 1.14 to 404.72 cM, with an estimated average length of 52.64 cM. The greatest distance between two adjacent markers was 48.25 cM. The scIvana_1-based markers (56) were positioned on 21 CGs, but were not regularly distributed. Interestingly, the distance between adjacent scIvana_1-based markers was less than 5 cM, and was observed on five CGs, suggesting a clustered organization. CONCLUSIONS: Results indicated the use of a NBS-profiling technique was efficient to develop retrotransposon-based markers in sugarcane. The simultaneous maximum-likelihood estimates of linkage and linkage phase based strategies confirmed the suitability of its approach to estimate linkage, and construct the linkage map. Interestingly, using our genetic data it was possible to calculate the number of retrotransposon scIvana_1 (~60) copies in the sugarcane genome, confirming previously reported molecular results. In addition, this research possibly will have indirect implications in crop economics e.g., productivity enhancement via QTL studies, as the mapping population parents differ in response to an important fungal disease.

Bax and Bcl-2 expression and TUNEL labeling in lumbar enlargement of neonatal rats after sciatic axotomy and melatonin treatment.

Peripheral axotomy in neonatal rats induces neuronal death. We studied the anti-apoptotic protein Bcl-2 and cell death promoter Bax in spinal cord of neonatal rats after sciatic transection and treatment with melatonin, a neuroprotective substance. Pups were unilaterally axotomized at P2 and received melatonin (1 mg/kg; sc) or vehicle 1 h prior to lesion, immediately after, at 1 h, 2 h and then once daily. Rats were sacrificed at 3 h, 6 h, 24 h, 72 h and 5 days postaxotomy. Intact animals were used as controls. Lumbar enlargement was processed for Nissl staining, immunohistochemistry and RT-PCR for Bax or Bcl-2 and TUNEL reaction. Motoneurons (MN) of lesioned (L) and normal (N) sides were counted, and MN survival ratio (MSR=L/N) was calculated. Bax and Bcl-2 showed cytoplasmic immunoreactivity (IR). Bax IR was noticeable in small cells but less evident in MN. In unlesioned pups, some Bax-positive small cells (B+) and TUNEL-positive nuclei (T+) were mainly seen in the dorsal horn. In lesioned animals given vehicle, Bax mRNA levels and numbers of B+ and T+ were increased in comparison with intact controls at 24 h postaxotomy. The basal IR for Bax in MN was not changed by axotomy. Bcl-2 IR was noted in all cells and, like Bcl-2 mRNA, was unaltered after lesion. Melatonin reduced MN loss at 24 h, 72 h and 5 days and T+ at 24 h after lesion but did not interfere with Bax or Bcl-2 expression. These results suggest that (1) sciatic transection at P2 increases Bax mRNA and the amount of B+ and T+ in the lumbar enlargement, (2) Bax IR in immature MN is not altered by axotomy and (3) melatonin protects MN and dorsal horn cells through a mechanism independent of Bax and Bcl-2.

The complete nucleotide sequence and genomic organization of Citrus Leprosis associated Virus, Cytoplasmatic type (CiLV-C).

The Citrus leprosis disease (CiL) is associated to a virus (CiLV) transmitted by Brevipalpus spp. mites (Acari: Tenuipalpidae). CiL is endemic in Brazil and its recently spreading to Central America represents a threat to citrus industry in the USA. Electron microscopy images show two forms of CiLV: a rare nuclear form, characterized by rod-shaped naked particle (CiLV-N) and a common cytoplasmic form (CiLV-C) associated with bacilliform-enveloped particle and cytoplasmic viroplasm. Due to this morphological feature, CiLV-C has been treated as Rhabdovirus-like. In this paper we present the complete nucleotide sequence and genomic organization of CiLV-C. It is a bipartite virus with sequence similarity to ssRNA positive plant virus. RNA1 encodes a putative replicase polyprotein and an ORF with no known function. RNA2 encodes 4 ORFs. pl5, p24 and p61 have no significant similarity to any known proteins and p32 encodes a protein with similarity to a viral movement protein. The CiLV-C sequences are associated with typical symptoms of CiL by RT-PCR. Phylogenetic analysis suggests that CiLV-C is probably a member of a new family of plant virus evolutionarily related to Tobamovirus.

Identification and genomic characterization of a new virus (Tymoviridae family) associated with citrus sudden death disease.

Citrus sudden death (CSD) is a new disease that has killed approximately 1 million orange trees in Brazil. Here we report the identification of a new virus associated with the disease. RNAs isolated from CSD-affected and nonaffected trees were used to construct cDNA libraries. A set of viral sequences present exclusively in libraries of CSD-affected trees was used to obtain the complete genome sequence of the new virus. Phylogenetic analysis revealed that this virus is a new member of the genus Marafivirus. Antibodies raised against the putative viral coat proteins allowed detection of viral antigens of expected sizes in affected plants. Electron microscopy of purified virus confirmed the presence of typical isometric Marafivirus particles. The screening of 773 affected and nonaffected citrus trees for the presence of the virus showed a 99.7% correlation between disease symptoms and the presence of the virus. We also detected the virus in aphids feeding on affected trees. These results suggest that this virus is likely to be the causative agent of CSD. The virus was named Citrus sudden death-associated virus.