Characterization of the Spodoptera littoralis nucleopolyhedrovirus type B lef-3 gene
| dc.contributor.author | Wolff, Jose Luiz Caldas | |
| dc.contributor.supervisor | Levin, David Bernard | |
| dc.date.accessioned | 2017-06-05T17:26:05Z | |
| dc.date.available | 2017-06-05T17:26:05Z | |
| dc.date.copyright | 1996 | en_US |
| dc.date.issued | 2017-06-05 | |
| dc.degree.department | Department of Biology | |
| dc.degree.level | Doctor of Philosophy Ph.D. | en_US |
| dc.description.abstract | We constructed a cDNA library with mRNA isolated from Sf9 cells infected with Spodoptera littoralis nucleopolyhedrovirus type B (SpliNPV-B) and identified the lef-3 gene from this library. Northern blot analysis showed that SpliNPV-B lef-3 mRNA was expressed as a 1.6 Kb transcript at 5 hours post infection (p.i.), reached high levels at 24 hours p.i., and remained highly expressed at 56 hours p.i.. Transcriptional mapping showed that lef-3 transcription started from two initiation sites (the distal and the proximal transcription initiation sites) located approximately 9 nucleotides apart. The sequences that modulate lef-3 expression were investigated by transient expression assays using a reporter gene under transcriptional control of the lef-3 promoter. Deletion analysis of the 5'-flanking region demonstrated that sequences up to 584 bases 5' of the distal transcription initiation site affected the level of reporter activity, indicating that this region contains transcription regulators. A region that was sufficient to direct basal level of promoter activity, the minimal promoter, was identified. This region encompasses the two transcription initiation sites, two TATA boxes, and a GATA motif. Mutations in the GATA motif resulted in substantial decrease in the level of reporter activity, suggesting that the GATA motif is an important element in the regulation of lef-3 gene expression. The sequence of a 2.6-kb region (mu 42.8-46.8) encompassing the lef-3 gene and flanking sequences was determined. Alignment of the predicted amino acid sequence of the LEF-3 polypeptide of SpliNPV-B with the putative sequences of AcMNPV and OpMNPV LEF-3 revealed low levels of sequence conservation (26% and 21% amino acid sequence identity, respectively). This low level of sequence conservation corroborates the view that, within the genus Nucleopolyhedrovirus, SpliNPV-B is distantly related to AcMNPV and OpMNPV. | en_US |
| dc.description.scholarlevel | Graduate | en_US |
| dc.identifier.uri | http://hdl.handle.net/1828/8225 | |
| dc.language | English | eng |
| dc.language.iso | en | en_US |
| dc.rights | Available to the World Wide Web | en_US |
| dc.subject | Spodoptera littoralis | en_US |
| dc.subject | Plants | en_US |
| dc.subject | Molecular biology | en_US |
| dc.title | Characterization of the Spodoptera littoralis nucleopolyhedrovirus type B lef-3 gene | en_US |
| dc.type | Thesis | en_US |