Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures

There are issues such as long maturation period, variable starch quality and lack of quality planting material that makes investors have less interest in commercial scale sago planting. Genetic transformation system for sago palm has the potential to improve the characteristic which would maximiz...

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Main Author:	Evra Raunie, Ibrahim
Format:	Thesis
Language:	English
Published:	2013
Subjects:	S Agriculture (General)
Online Access:	http://ir.unimas.my/id/eprint/8789/1/Evra%20Raunie%20Ibrahim%20ft.pdf
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spelling	my-unimas-ir.87892023-10-03T04:05:52Z Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures 2013 Evra Raunie, Ibrahim S Agriculture (General) There are issues such as long maturation period, variable starch quality and lack of quality planting material that makes investors have less interest in commercial scale sago planting. Genetic transformation system for sago palm has the potential to improve the characteristic which would maximized yield per Ha and subsequently attract more investors. Therefore this study was aimed to optimized sago genetic transformation system in order to improve the characteristic of existing sago palm and to develop a suspension culture system to support mass production of sago clonal planting material. This study consists of development of sago palm suspension culture, determination of minimal inhibitory concentration of Basta™ on sago, development of sago transformation system via Agrobacterium and gene gun. The suspension culture of sago palm was successfully propagated using 4.5-15 gram of inoculum in every litre of media containing 0.18-1.8 mg/l NAA and 1-1.5 mg/l 2, 4-D. The propagated cells are ready for transformation after 3 months cycle. The minimal inhibitory concentration of Basta™ for sago palm was determined at 30 mg/l. The genetic transformation of sago target tissues derived from suspension culture was conducted using two methods; the Agrobacterium tumefaciens strain LB4404 containing plasmid vector pGSA1131 and the Biorad™ Helios Gene Gun System. The putative transgenic embryoids were selected through Basta™ selection process using media containing minimum concentration of Basta™ at 30 mg/l. The presence of bar and gus genes in selected regenerated callus were verified by PCR amplification, GUS staining analysis and dot blot analysis. From this study it is proven that iv the most suitable plant material for transformation is the embryogenic callus (D0E/EC) compared to embryoid stage (D1 and D2) and early formation of plantlets (D3) for both methods. As for the high sugar treatment during and after infection, the wounding using gold particle bombardment and the sonication treatment on the target cells had also proven able to increase the efficiency of the transformation via Agrobacterium. The transformation using Helios Gene Gun showed a higher transformation rates when target tissues at embryogenic stage (D0E/EC) were bombarded with 280psi of helium pressure at 6 to 8 cm of distance and with the number of firing once and twice of bombardment. Universiti Malaysia Sarawak, (UNIMAS) 2013 Thesis http://ir.unimas.my/id/eprint/8789/ http://ir.unimas.my/id/eprint/8789/1/Evra%20Raunie%20Ibrahim%20ft.pdf text en validuser masters Universiti Malaysia Sarawak, (UNIMAS) Faculty of Resource Science and Technology
institution	Universiti Malaysia Sarawak
collection	UNIMAS Institutional Repository
language	English
topic	S Agriculture (General)
spellingShingle	S Agriculture (General) Evra Raunie, Ibrahim Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
description	There are issues such as long maturation period, variable starch quality and lack of quality planting material that makes investors have less interest in commercial scale sago planting. Genetic transformation system for sago palm has the potential to improve the characteristic which would maximized yield per Ha and subsequently attract more investors. Therefore this study was aimed to optimized sago genetic transformation system in order to improve the characteristic of existing sago palm and to develop a suspension culture system to support mass production of sago clonal planting material. This study consists of development of sago palm suspension culture, determination of minimal inhibitory concentration of Basta™ on sago, development of sago transformation system via Agrobacterium and gene gun. The suspension culture of sago palm was successfully propagated using 4.5-15 gram of inoculum in every litre of media containing 0.18-1.8 mg/l NAA and 1-1.5 mg/l 2, 4-D. The propagated cells are ready for transformation after 3 months cycle. The minimal inhibitory concentration of Basta™ for sago palm was determined at 30 mg/l. The genetic transformation of sago target tissues derived from suspension culture was conducted using two methods; the Agrobacterium tumefaciens strain LB4404 containing plasmid vector pGSA1131 and the Biorad™ Helios Gene Gun System. The putative transgenic embryoids were selected through Basta™ selection process using media containing minimum concentration of Basta™ at 30 mg/l. The presence of bar and gus genes in selected regenerated callus were verified by PCR amplification, GUS staining analysis and dot blot analysis. From this study it is proven that iv the most suitable plant material for transformation is the embryogenic callus (D0E/EC) compared to embryoid stage (D1 and D2) and early formation of plantlets (D3) for both methods. As for the high sugar treatment during and after infection, the wounding using gold particle bombardment and the sonication treatment on the target cells had also proven able to increase the efficiency of the transformation via Agrobacterium. The transformation using Helios Gene Gun showed a higher transformation rates when target tissues at embryogenic stage (D0E/EC) were bombarded with 280psi of helium pressure at 6 to 8 cm of distance and with the number of firing once and twice of bombardment.
format	Thesis
qualification_level	Master's degree
author	Evra Raunie, Ibrahim
author_facet	Evra Raunie, Ibrahim
author_sort	Evra Raunie, Ibrahim
title	Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
title_short	Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
title_full	Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
title_fullStr	Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
title_full_unstemmed	Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
title_sort	development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures
granting_institution	Universiti Malaysia Sarawak, (UNIMAS)
granting_department	Faculty of Resource Science and Technology
publishDate	2013
url	http://ir.unimas.my/id/eprint/8789/1/Evra%20Raunie%20Ibrahim%20ft.pdf
_version_	1783727993657491456

Development of sago suspension culture system and optimization of transformation system for sago palm (metroxylon sagu rottb.) cultures

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