Isolation of the SEC6 cDNA Sequence from the Black Widow Spider
Poster Number
11
Format
Poster Presentation
Faculty Mentor Name
Craig Vierra
Abstract/Artist Statement
Spider silk has extraordinary mechanical properties and scientists have been attempting to produce synthetic silk for commercial applications, which include usage for bullet proof vests, ropes and cords, sutures and aircraft wings. Previously our lab identified several key target genes that are linked to the silking process in black widow spiders. Using DNA microaiTay analysis, we identified a few genes that had their expression profiles change upon pulling dragline silk from black widow spiders. One of these genes, which represented a partial eDNA sequence, showed similarity to the yeast secretory protein SEC6. This similarity, which was determined by searching the NCBI nr nucleic acid database against our partial eDNA sequence, supports this gene represents the orthologue of yeast sec6 and may play a role in the assembly process of silks during extrusion. In these studies, our goal was to isolate the full-length eDNA sequence of black widow sec6. To amplify the complete eDNA sequence, we employed 5' and 3' anchored PCR, which are forms of rapid amplification of eDNA ends (5' and 3' RACE). Anchored PCR led to the successful amplification of the 5' as well as the 3' end of the sec6 gene. Following the amplification of the eDNA inserts, these pieces of nucleic acid were placed into a cloning vector and sequenced. The full-length sequence was reconstructed using overlapping pieces of the nucleic acid pieces obtained by RACE. Our long-term goal is to be able to overexpress black widow SEC6 in bacteria, generate antibodies against SEC6 in rabbits, and use the antibodies to sttidy the protein in vivo to learn more regarding its potential role in the extrusion process of silks in spiders.
Location
Wendell Phillips Center, 1st floor hallways
Start Date
3-5-2008 1:00 PM
End Date
3-5-2008 3:00 PM
Isolation of the SEC6 cDNA Sequence from the Black Widow Spider
Wendell Phillips Center, 1st floor hallways
Spider silk has extraordinary mechanical properties and scientists have been attempting to produce synthetic silk for commercial applications, which include usage for bullet proof vests, ropes and cords, sutures and aircraft wings. Previously our lab identified several key target genes that are linked to the silking process in black widow spiders. Using DNA microaiTay analysis, we identified a few genes that had their expression profiles change upon pulling dragline silk from black widow spiders. One of these genes, which represented a partial eDNA sequence, showed similarity to the yeast secretory protein SEC6. This similarity, which was determined by searching the NCBI nr nucleic acid database against our partial eDNA sequence, supports this gene represents the orthologue of yeast sec6 and may play a role in the assembly process of silks during extrusion. In these studies, our goal was to isolate the full-length eDNA sequence of black widow sec6. To amplify the complete eDNA sequence, we employed 5' and 3' anchored PCR, which are forms of rapid amplification of eDNA ends (5' and 3' RACE). Anchored PCR led to the successful amplification of the 5' as well as the 3' end of the sec6 gene. Following the amplification of the eDNA inserts, these pieces of nucleic acid were placed into a cloning vector and sequenced. The full-length sequence was reconstructed using overlapping pieces of the nucleic acid pieces obtained by RACE. Our long-term goal is to be able to overexpress black widow SEC6 in bacteria, generate antibodies against SEC6 in rabbits, and use the antibodies to sttidy the protein in vivo to learn more regarding its potential role in the extrusion process of silks in spiders.