2026-09-26T01:49:16Zhttps://mro.massey.ac.nz/server/oai/requestoai:mro.massey.ac.nz:10179/43192025-11-11T14:08:23Zcom_10179_294col_10179_296
DSpace at Massey University
author
Yang, Tian
2013-05-03T00:20:52Z
2013-05-03T00:20:52Z
2013
http://hdl.handle.net/10179/4319
Q112901980
https://www.wikidata.org/wiki/Q112901980
Lentiviral vectors, originally derived from human immunodeficiency virus, provide highly efficient viral gene delivery vehicles. Lentiviral vectors often use a constitutive promoter to drive the expression of a therapeutic gene. To regulate the expression of a therapeutic gene, a regulatory system such as Tet-On needs to be established in the target cell lines to produce a regulatory protein, reverse Tet-responsive transcriptional activator (rtTA). The expressed rtTA binds to the tetracycline responsive element (TRE) in the promoter in response to doxycycline and activates transcription of gene of interest. A hypothesis in this study is based on the speculation that a basal leaky expression of rtTA in the bi-directional TRE vectors allows instantly inducible expression of a gene of interest and thereby avoids the time-consuming procedures for generating Tet-On cell lines. Based on this hypothesis, a novel lentiviral vector has been developed to examine an instant induction of PP2Cβ as a target gene. Three instantly inducible bicistronic lentiviral vectors [pLenti-Bi-TRE-Tet-on (V), pLenti-Bi-TRE-Tet-on-PP2Cβ WT (WT), pLenti- Bi-TRE-Tet-on-PP2Cβ MUT (MUT)] were constructed and characterised to assess the usefulness of these vectors. Transient transfection of both WT and MUT vectors into HEK293T cells showed a great induction of PP2Cβ expression upon 24 h of 1 μM doxycycline treatment. The result promises the use of these vectors as a mammalian expression plasmid with a feature of inducible target gene expression. However, viral infection studies involving lentiviral packaging and infection procedures did not show a reproducible expression of rtTA or PP2Cβ in HEK293T cells. Therefore, the inducibility of viral transduction needs to be improved for the future studies of PP2Cβ in primary cells.
en
The Author
Lentiviral vectors
Gene therapy
Gene expression
Gene regulation
Tetracycline
Genetic vectors
Development of a tetracycline-inducible lentiviral vector with an instant regulatory system : a thesis presented in partial fulfillment of the requirements for the degree of Master of Science (MSc) in Biochemistry at Massey University, Manawatu, New Zealand
Thesis
PGgyPkNPUFlSSUdIVDwvaDI+DQo8cD5Db3B5cmlnaHQgcmVzaWRlcyB3aXRoIHRoZSBhdXRob3IsIGFuZCBpcyBnb3Zlcm5lZCBieSB0aGUgQ29weXJpZ2h0IEFjdCAxOTk0LiANCg0KPHA+VW5sZXNzIHlvdSBzdGF0ZSBvdGhlcndpc2UsIHRoZSANCkxpYnJhcnkgbWF5IGNvcHkgYWxsIG9yIHBhcnQgb2YgeW91ciB0aGVzaXMgPGJyPg0KYW5kIHByb3ZpZGUgaXQgdG8gYW5vdGhlciBpbnN0aXR1dGlvbiBmb3IgdGhlIHB1cnBvc2VzIG9mIHJlc2VhcmNoIGFuZCBwcml2YXRlIHN0dWR5LjwvcD4NCg0KPHA+WW91IG1heSBhc3NlcnQgeW91ciBtb3JhbCByaWdodCB0byBiZSBpZGVudGlmaWVkIGFzIHRoZSBhdXRob3Igb2YgdGhlIHdvcmsgaW4gYSBzdGF0ZW1lbnQgdG8gdGhpcyA8YnI+ZWZmZWN0IG9uIA0KdGhlIHZlcnNpb24gb2YgdGhlIHRpdGxlIHBhZ2UuPC9wPg0KDQo8cD5Nb3JlIGluZm9ybWF0aW9uIGNhbiBiZSBvYnRhaW5lZCBmcm9tIE1hc3NleSBVbml2ZXJzaXR5J3MgPGEgIGhyZWY9Imh0dHA6Ly9jb3B5cmlnaHQubWFzc2V5LmFjLm56LyI+Q29weXJpZ2h0IA0KDQpJbmZvcm1hdGlvbjwvYT4gd2ViIHNpdGUuIDwvcD4NCjxwPiBZb3Ugc2hvdWxkIGFsc28gYmUgYXdhcmUgb2YgTWFzc2V5IFVuaXZlcnNpdHkncyA8YSANCg0KaHJlZj0iaHR0cDovL3BvbGljeWd1aWRlLm1hc3NleS5hYy5uei9tYXNzZXkvZm9yL3N0YWZmL3BvbGljeS9wb2xpY2llcy9yZXNlYXJjaC9pbnRlbGxlY3R1YWwtcHJvcGVydHktDQoNCnBvbGljeS5jZm0iPkludGVsbGVjdHVhbCBQcm9wZXJ0eSBQb2xpY3k8L2E+LiA8L3A+
URL
https://mro.massey.ac.nz/bitstreams/59a974df-466f-4b89-859c-01baa3c81a0e/download
File
MD5
3ef8ef97c393f63e367830aaf8139f21
2153308
application/pdf
02_whole.pdf
URL
https://mro.massey.ac.nz/bitstreams/7af52c63-64f3-4f3c-8fa5-b85cac306a22/download
File
MD5
9b70a1a562ad085995d3524ca5d07bd1
333492
application/pdf
01_front.pdf
URL
https://mro.massey.ac.nz/bitstreams/61dddfb0-81a1-40d8-b407-70fc74ba75c6/download
File
MD5
b34eea43e88fdfbf640d38b2dd2d0df8
20182
text/plain
01_front.pdf.txt
URL
https://mro.massey.ac.nz/bitstreams/a933d15f-e2cc-44de-b6ca-841dd3f78d35/download
File
MD5
934cc52718e3e9ea7468ecbf8f64415b
201188
text/plain
02_whole.pdf.txt