Polydnavirus delivery constructs
View Patent ↗Provided herein are methods of producing a genetically modified cell by introducing a polydnavirus delivery construct to a target cell. The polydnavirus delivery construct can comprise an exogenous nucleic acid to form a genetically modified cell comprising the exogenous nucleic acid. Also provided are polydnavirus delivery constructs comprising an exogenous nucleic acid, as well as polydnavirus virions and genetically modified cells comprising the same. Further provided are in vitro methods of identifying a transformed cell. The methods comprise introducing a vector comprising a nucleotide sequence encoding a glc polypeptide to an adherent cell and cultivating the cell under conditions that allow for the expression of the glc polypeptide. Expression of the glc polypeptide results in a transformed cell that is identified by a loss of adherency.
1. A method of integrating exogenous DNA into the genome of a cell thereby producing a genetically modified cell, comprising introducing a polydnavirus delivery construct comprising an exogenous nucleic acid to a target cell to form the genetically modified cell comprising the exogenous nucleic acid, wherein the polydnavirus delivery construct is derived from Microplitis demolitor bracovirus (MdBV).
2. The method of claim 1 , further comprising introducing to the target cell an integrase or a nucleic acid encoding an integrase.
3. The method of claim 2 , wherein the integrase is a proviral bracovirus integrase.
4. The method of claim 1 , wherein the polydnavirus delivery construct is encapsidated with a polydnavirus virion, wherein the method comprises introducing the polydnavirus virion to the cell.
5. The method of claim 4 , wherein the polydnavirus virion further comprises the nucleic acid encoding the integrase.
6. The method of claim 1 , comprising introducing the polydnavirus delivery construct to the cell by transfection or electroporation.
7. The method of claim 1 , wherein the polydnavirus comprises at least one host integration motif.
8. The method of claim 7 , wherein the host integration motif is selected from the group consisting of SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, and SEQ ID NO:43.
9. The method of claim 7 , wherein the host integration motif is comprised within an MdBV segment.
10. The method of claim 9 , wherein the MdBV segment is selected from the group consisting of segment A, segment B, segment C, segment D, segment E, segment F, segment G, segment H, segment I, segment J, segment K, segment L, segment M, segment N, and segment O.
11. The method of claim 1 , wherein the exogenous nucleic acid comprises a nucleotide sequence of about 50 to about 35,000 nucleotides.
12. The method of claim 11 , wherein the nucleotide sequence is about 1,000 to about 20,000 nucleotides.
13. The method of claim 1 , wherein the exogenous nucleic acid sequence encodes a recombinant polypeptide.
14. The method of claim 13 , wherein the recombinant polypeptide comprises a detectable tag.
15. The method of claim 14 , wherein the detectable tag is selected from the group consisting of a histidine tag, a FLAG tag, a GST tag, and a hemagglutinin tag.