Construction of a vector containing hygromycin (HPT) gene driven by double 35S (2XCaMV35S) promoter for oil palm transformation

Bahariah Bohari, . and Abdul Masani Mat Yunus, . and Omar Abdul Rasid, . and Ahmad Parveez Ghulam Kadir, . Construction of a vector containing hygromycin (HPT) gene driven by double 35S (2XCaMV35S) promoter for oil palm transformation. pp. 180-188. ISSN 1511-2780

Transformation vector construction is one of the important disciplines for plant genetic transformation studies. A series of vector consisting of hygromycin phosphotransferase (hpt) gene as the selective marker and green fluorescent protein (GFP) as the visual reporter gene under the control of double cauliflower mosaic virus 35S (2XCaMV35S) promoter has been engineered for transformation into oil palm cells. These genes were cloned into different types of cloning and expression vectors. The cloning was carried out by using restriction enzyme digestion and ligation method. Five intermediate vectors have been created for insertion of 2XCaMV35S-HPT-35ST and 2XCaMV35S-GFP-35ST into modified pBINPLUS backbone vector for particle bombardment and Agrobacterium-based transformation protocols. All vectors were sequenced to confirm the integrity of DNA region. The vectors were later transformed into oil palm embryogenic calli using biolistic device. The viability of the vectors was initially evaluated by transient GFP fluorescence expression observed under fluorescence microscope. It was demonstrated that the 2XCaMV35S promoter was able to drive the expression of gfp as gene in oil palm calli.

Full text available from: Official URL: http://jopr.mpob.gov.my/construction-of-a-vector-c...

Abstract

Transformation vector construction is one of the important disciplines for plant genetic transformation studies. A series of vector consisting of hygromycin phosphotransferase (hpt) gene as the selective marker and green fluorescent protein (GFP) as the visual reporter gene under the control of double cauliflower mosaic virus 35S (2XCaMV35S) promoter has been engineered for transformation into oil palm cells. These genes were cloned into different types of cloning and expression vectors. The cloning was carried out by using restriction enzyme digestion and ligation method. Five intermediate vectors have been created for insertion of 2XCaMV35S-HPT-35ST and 2XCaMV35S-GFP-35ST into modified pBINPLUS backbone vector for particle bombardment and Agrobacterium-based transformation protocols. All vectors were sequenced to confirm the integrity of DNA region. The vectors were later transformed into oil palm embryogenic calli using biolistic device. The viability of the vectors was initially evaluated by transient GFP fluorescence expression observed under fluorescence microscope. It was demonstrated that the 2XCaMV35S promoter was able to drive the expression of gfp as gene in oil palm calli.

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Item Type:	Article
AGROVOC Term:	Elaeis guineensis
AGROVOC Term:	Oil palms
AGROVOC Term:	Genetic vectors
AGROVOC Term:	DNA cloning
AGROVOC Term:	Gene expression
AGROVOC Term:	Genes
AGROVOC Term:	Genetic engineering
AGROVOC Term:	Genetic transformation
AGROVOC Term:	Genetically engineered organisms
AGROVOC Term:	Polymerase chain reaction
Geographical Term:	Malaysia
Depositing User:	Ms. Suzila Mohamad Kasim
Last Modified:	28 Apr 2025 07:07
URI:	http://webagris.upm.edu.my/id/eprint/24195

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