Bacterial Expression of Fasciclin: A Key Component of Dragline Silk
Poster Number
02B
Format
Poster Presentation
Faculty Mentor Name
Craig Vierra
Faculty Mentor Department
Biological Sciences
Abstract/Artist Statement
Dragline silk, the primary locomotive silk produced by the black widow spider, can be used for a number of industrial applications, due to its tensile strength, extensibility, and toughness. Its material properties give it considerable versatility, since it has greater tensile strength than high tensile steel and is comparable toughness to Kevlar (body armour). Furthermore, dragline silk is eco-friendly, consisting of organic properties that make it biodegradable. Upon proteomic analysis, dragline silk was found to be composed of 7 different proteins common to the major ampullate gland, the spinning dope, and dragline silk. These proteins include MaSp1, MaSp2, CRISP3, fasciclin, and 3 cysteine rich proteins: CRP1, CRP2, CRP4. This study focused primarily on fasciclin, one of the 7 common proteins. Isolation of the fasciclin cDNA was accomplished by PCR amplification of the gene. The resulting fasciclin cDNA was amplified, extracted via agarose gel electrophoresis, then inserted into a pET-19b SUMO plasmid expression vector. Next, we transformed the expression vector into competent E.coli cells for expression studies. Following purification of fasciclin, mass spectroscopy was used to validate the purification of the protein after affinity purification. Our results demonstrate that we successfully expressed and purified the fasciclin protein from bacterial cells.
Location
DeRosa University Center, Ballroom
Start Date
29-4-2017 1:00 PM
End Date
29-4-2017 3:00 PM
Bacterial Expression of Fasciclin: A Key Component of Dragline Silk
DeRosa University Center, Ballroom
Dragline silk, the primary locomotive silk produced by the black widow spider, can be used for a number of industrial applications, due to its tensile strength, extensibility, and toughness. Its material properties give it considerable versatility, since it has greater tensile strength than high tensile steel and is comparable toughness to Kevlar (body armour). Furthermore, dragline silk is eco-friendly, consisting of organic properties that make it biodegradable. Upon proteomic analysis, dragline silk was found to be composed of 7 different proteins common to the major ampullate gland, the spinning dope, and dragline silk. These proteins include MaSp1, MaSp2, CRISP3, fasciclin, and 3 cysteine rich proteins: CRP1, CRP2, CRP4. This study focused primarily on fasciclin, one of the 7 common proteins. Isolation of the fasciclin cDNA was accomplished by PCR amplification of the gene. The resulting fasciclin cDNA was amplified, extracted via agarose gel electrophoresis, then inserted into a pET-19b SUMO plasmid expression vector. Next, we transformed the expression vector into competent E.coli cells for expression studies. Following purification of fasciclin, mass spectroscopy was used to validate the purification of the protein after affinity purification. Our results demonstrate that we successfully expressed and purified the fasciclin protein from bacterial cells.