Full Text Available
Note: Clicking the button above will open the full text document at the original institutional repository in a new window.
The expression of Dianthin 30, a ribosome inactivating protein
Thesis (MSc)--Stellenbosch University, 2003.
Saved in:
| Main Author: | |
|---|---|
| Other Authors: | |
| Format: | Thesis |
| Language: | en_ZA |
| Published: |
Stellenbosch : Stellenbosch University
2012
|
| Subjects: | |
| Tags: |
No Tags, Be the first to tag this record!
|
| _version_ | 1867614070245425152 |
|---|---|
| access_status_str | Open Access |
| author | Maree, H. J. (Hans Jacob) |
| author2 | Burger, Johan T. |
| author_browse | Burger, Johan T. Maree, H. J. (Hans Jacob) |
| author_facet | Burger, Johan T. Maree, H. J. (Hans Jacob) |
| author_sort | Maree, H. J. (Hans Jacob) |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MSc)--Stellenbosch University, 2003. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/53633 |
| institution | Stellenbosch University (South Africa) |
| language | en_ZA |
| last_indexed | 2026-06-10T12:46:11.731Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2012 |
| publishDateRange | 2012 |
| publishDateSort | 2012 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/53633 The expression of Dianthin 30, a ribosome inactivating protein Maree, H. J. (Hans Jacob) Burger, Johan T. Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics. Institute for Plant Biotechnology (IPB) Plants -- Virus resistance -- Genetic aspects Virus diseases of plants Plant viruses -- Control Transgenic plants Thesis (MSc)--Stellenbosch University, 2003. ENGLISH ABSTRACT: Ribosome inactivating proteins (RIPs) are currently classified as rRNA N-glycosidases, but also have polynucleotide: adenosine glycosidase activity. RIPs are believed to have anti-viral and anti-fungal properties, but the exact mechanism of these proteins still need to be elucidated.The mechanism of resistance however, appears to be independent of the pathogen. For resistance the RIP terminates virus infected plant cells and stops the reproduction and spread of the virus. Transgenic plants containing RIPs should thus be resistant to a wide range of viruses. The ultimate goal of the larger project of which this forms part is the development of virus resistant plants. To monitor the expression of a RIP in a transgenic plant a detection method had to be developed. Antibody detection of the RIP was decided upon as the most cost effective method. The RIP, Dianthin 30 from Dianthus caryophyllus (carnation), was used and expressed in bacterial and insect expression systems. The bacterial expression experiments were done using the pET expression system in BL21(DE3)pLysS cells. The expression in this system yielded recombinant protein at a very low concentration. Expression experiments were also performed in insect tissue culture with the baculovirus vector BAC-TO-BAC™.With this system the expression was also too low to be used for the production of antibodies. A Dianthin 30 specific peptide was then designed and then produced by Bio-Synthesis. This peptide was then used to raise antibodies to detect Dianthin 30. These antibodies were tested on Dianthus caryophyllus proteins. To establish if this detection method was effective to monitor the expression in plants, tobacco plants were transformed with Agrobacterium tumefaciens containing Dianthin 30 in the pART27 plant expression vector. The putative transformed plants were analysed with peR and Southern blots. AFRIKAANSE OPSOMMING: Tans word Ribosomale-inaktiverende proteïene (RIPs) geklassifiseer as rRNA N-glikosidase wat ook polinukleotied: adenosien glikosidase aktiwiteit bevat. Daar word geglo dat RIPs anti-virale en anti-fungus eienskappe bevat, maar die meganisme van beskerming word nog nie ten volle verstaan nie. Dit is wel bewys dat die meganisme van weerstand onafhanklik is van die patogeen. Virus geinfekteerde plantselle word deur die RIP gedood om die voortplanting en verspreiding te bekamp en sodoende word weerstand bewerkstellig. Transgeniese plante wat dan 'n RIP bevat sal dus weerstandbiedend wees teen 'n wye spektrum virusse. Die hoofdoel van die breër projek, waarvan die projek deel uitmaak: is die ontwikkeling van virusbestande plante. Om die uitdrukking van die RIP in die transgeniese plante te kontroleer, moes 'n deteksie metode ontwikkel word. Die mees koste effektiewe deteksie metode is met teenliggame. Die RIP, Dianthin 30 from Dianthus caryophyllus (angelier) was gebruik vir uitdrukking in bakteriele- en insekweefselkultuur. Die bakteriele uitdrukkingseksperimente was gedoen met die pET uitdrukkings sisteem III BL21(DE3)pLysS selle. Die uitdrukking in die sisteem het slegs rekombinante proteïene gelewer in uiters lae konsentrasies. Uitdrukkingseksperimente was ook gedoen in insekweefselkultuur met die baculovirus vektor BAC-To- BACTM. Met die sisteem was die uitdrukking ook veels te laag om bruikbaar te wees vir die produksie van teenliggame. Daar is toe 'n peptied ontwerp wat Dianthin 30 kan verteenwoordig vir die produksie van teenliggame. Die teenliggame is getoets teen Dianthus caryophyllus proteïene. Om vas te stel of die deteksiemetode wel die uitdrukking van Dianthin 30 sal kan monitor, is tabak ook getransformeer met Dianthin 30. Die transformasies is gedoen met die hulp van Agrobacterium tumefaciens en die pART27 plant uitdrukkings vektor. Die plante is getoets met die polimerase ketting reaksie en Southern klad tegnieke. Masters 2012-08-27T11:35:34Z 2012-08-27T11:35:34Z 2003-03 Thesis http://hdl.handle.net/10019.1/53633 en_ZA Stellenbosch University 91 pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Plants -- Virus resistance -- Genetic aspects Virus diseases of plants Plant viruses -- Control Transgenic plants Maree, H. J. (Hans Jacob) The expression of Dianthin 30, a ribosome inactivating protein |
| title | The expression of Dianthin 30, a ribosome inactivating protein |
| title_full | The expression of Dianthin 30, a ribosome inactivating protein |
| title_fullStr | The expression of Dianthin 30, a ribosome inactivating protein |
| title_full_unstemmed | The expression of Dianthin 30, a ribosome inactivating protein |
| title_short | The expression of Dianthin 30, a ribosome inactivating protein |
| title_sort | expression of dianthin 30 a ribosome inactivating protein |
| topic | Plants -- Virus resistance -- Genetic aspects Virus diseases of plants Plant viruses -- Control Transgenic plants |
| url | http://hdl.handle.net/10019.1/53633 |
| work_keys_str_mv | AT mareehjhansjacob theexpressionofdianthin30aribosomeinactivatingprotein AT mareehjhansjacob expressionofdianthin30aribosomeinactivatingprotein |