ERROR LOADING HTML FROM SOURCE (http://ncf.sobek.ufl.edu//design/skins/UFDC/html/header_item.html)

Baculovirus Overexpression of RNA Helicase A from C. elegens and Bandshift Analysis of Its RNA Binding Properties

Permanent Link: http://ncf.sobek.ufl.edu/NCFE003198/00001

Material Information

Title: Baculovirus Overexpression of RNA Helicase A from C. elegens and Bandshift Analysis of Its RNA Binding Properties
Physical Description: Book
Language: English
Creator: Batts, Shelley
Publisher: New College of Florida
Place of Publication: Sarasota, Fla.
Creation Date: 2003
Publication Date: 2003

Subjects

Subjects / Keywords: RNA
Helicase
C.elegens
Baculovirus
RNA Helicase A
Protein Binding
Genre: bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: The rha-1gene from C. elegans was inserted into a pFastBac donor plasmid and transformed into E. coli colonies. These colonies were subsequently screened for correct positioning of the recombinant DNA. Positive clones were recombined into bacmid DNA which was purified and used to infect Sf9 cells which will express the RHA- 1 (RNA Helicase A) protein. Protein expression was tested with Western blot analysis, and the infected cells which proved to be overexpressing RHA- 1 were lysed, and the RHA- 1 protein was purified on a Ni- NTA resin colunm. Fractions obtained from the purification were compared by SDS-PAGE and Western blot analysis, and the fractions containing the largest concentration of protein were dialyzed. The resultant purified, dialyzed RHA- 1 protein was then tested for binding with single- and double-stranded RNA using an Electrophoretic Mobility Shift Assay (EMSA). These assays were inconclusive and suggested little to no binding was occurring. Although multiple optimization experiments testing different variable were conducted, it can be assumed that the correct binding conditions were not found.
Statement of Responsibility: by Shelley Batts
Thesis: Thesis (B.A.) -- New College of Florida, 2003
Electronic Access: RESTRICTED TO NCF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE
Bibliography: Includes bibliographical references.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The New College of Florida, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Local: Faculty Sponsor: Walstrom, Katherine

Record Information

Source Institution: New College of Florida
Holding Location: New College of Florida
Rights Management: Applicable rights reserved.
Classification: local - S.T. 2003 B3
System ID: NCFE003198:00001

Permanent Link: http://ncf.sobek.ufl.edu/NCFE003198/00001

Material Information

Title: Baculovirus Overexpression of RNA Helicase A from C. elegens and Bandshift Analysis of Its RNA Binding Properties
Physical Description: Book
Language: English
Creator: Batts, Shelley
Publisher: New College of Florida
Place of Publication: Sarasota, Fla.
Creation Date: 2003
Publication Date: 2003

Subjects

Subjects / Keywords: RNA
Helicase
C.elegens
Baculovirus
RNA Helicase A
Protein Binding
Genre: bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: The rha-1gene from C. elegans was inserted into a pFastBac donor plasmid and transformed into E. coli colonies. These colonies were subsequently screened for correct positioning of the recombinant DNA. Positive clones were recombined into bacmid DNA which was purified and used to infect Sf9 cells which will express the RHA- 1 (RNA Helicase A) protein. Protein expression was tested with Western blot analysis, and the infected cells which proved to be overexpressing RHA- 1 were lysed, and the RHA- 1 protein was purified on a Ni- NTA resin colunm. Fractions obtained from the purification were compared by SDS-PAGE and Western blot analysis, and the fractions containing the largest concentration of protein were dialyzed. The resultant purified, dialyzed RHA- 1 protein was then tested for binding with single- and double-stranded RNA using an Electrophoretic Mobility Shift Assay (EMSA). These assays were inconclusive and suggested little to no binding was occurring. Although multiple optimization experiments testing different variable were conducted, it can be assumed that the correct binding conditions were not found.
Statement of Responsibility: by Shelley Batts
Thesis: Thesis (B.A.) -- New College of Florida, 2003
Electronic Access: RESTRICTED TO NCF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE
Bibliography: Includes bibliographical references.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The New College of Florida, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Local: Faculty Sponsor: Walstrom, Katherine

Record Information

Source Institution: New College of Florida
Holding Location: New College of Florida
Rights Management: Applicable rights reserved.
Classification: local - S.T. 2003 B3
System ID: NCFE003198:00001

ERROR LOADING HTML FROM SOURCE (http://ncf.sobek.ufl.edu//design/skins/UFDC/html/footer_item.html)