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Evaluation on the effectiveness of 2-deoxyglucose-6-phosphate phosphatase (DOG(R)1) gene as a selectable marker for oil palm (Elaeis guineensis Jacq.) embryogenic calli transformation mediated by Agrobacterium tumefaciens.

Izawati AM, Masani MY, Ismanizan I, Parveez GK - Front Plant Sci (2015)

Bottom Line: The plantlets were later transferred into soil and grown in a biosafety screenhouse.PCR and subsequently Southern blot analyses were carried out to confirm the integration of the transgene in the plantlets.This result demonstrates the potential of using combination of DOG(R)1 gene and 2-DOG for regenerating transgenic oil palm.

View Article: PubMed Central - PubMed

Affiliation: Advanced Biotechnology and Breeding Centre, Malaysian Palm Oil Board Selangor, Malaysia.

ABSTRACT
DOG(R)1, which encodes 2-deoxyglucose-6-phosphate phosphatase, has been used as a selectable marker gene to produce transgenic plants. In this study, a transformation vector, pBIDOG, which contains the DOG(R)1 gene, was transformed into oil palm embryogenic calli (EC) mediated by Agrobacterium tumefaciens strain LBA4404. Transformed EC were exposed to 400 mg l(-1) 2-deoxyglucose (2-DOG) as the selection agent. 2-DOG resistant tissues were regenerated into whole plantlets on various regeneration media containing the same concentration of 2-DOG. The plantlets were later transferred into soil and grown in a biosafety screenhouse. PCR and subsequently Southern blot analyses were carried out to confirm the integration of the transgene in the plantlets. A transformation efficiency of about 1.0% was obtained using DOG(R)1 gene into the genome of oil palm. This result demonstrates the potential of using combination of DOG(R)1 gene and 2-DOG for regenerating transgenic oil palm.

No MeSH data available.


Related in: MedlinePlus

Schematic diagram of DOGR1 expression cassette (line indicates the size of 1.5 kb) used as probe for Southern blot analysis. RB, right border; LB, left border; NOS, nopaline synthase gene terminator; DOGR1, gene codes for 2-deoxyglucose-6-phosphate phosphatase; CaMV35S, cauliflower mosaic virus 35S promoter. Arrows indicates the position of primers used for PCR analysis to amplify 741 bp.
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Figure 1: Schematic diagram of DOGR1 expression cassette (line indicates the size of 1.5 kb) used as probe for Southern blot analysis. RB, right border; LB, left border; NOS, nopaline synthase gene terminator; DOGR1, gene codes for 2-deoxyglucose-6-phosphate phosphatase; CaMV35S, cauliflower mosaic virus 35S promoter. Arrows indicates the position of primers used for PCR analysis to amplify 741 bp.

Mentions: The construction of pBIDOG plasmid was performed by replacing the DNA fragment of CaMV35S-GUS-Nos of pBI35SGUS with the DNA fragment of CaMV35S-DOGR1-Nos of pBINARDOG (SunGene, Germany). pBI121 plasmid was digested with PmeI and ClaI to remove the DNA fragment of NosPro-nptII-Nos, rendered blunt and religated to generate plasmid pBI35SGUS. The CaMV35S-DOGR1-Nos fragment was released from pBINARDOG by HindIII and EcoRI digestion, and cloned into pBI35SGUS releasing the DNA fragment of CaMV35S-GUS-Nos at similar sites to generate pBIDOG (Figure 1). pBIDOG was transformed into E. coli and later mobilized into Agrobacterium tumefaciens strain LBA4404 via electroporation and used as a vector for oil palm transformation.


Evaluation on the effectiveness of 2-deoxyglucose-6-phosphate phosphatase (DOG(R)1) gene as a selectable marker for oil palm (Elaeis guineensis Jacq.) embryogenic calli transformation mediated by Agrobacterium tumefaciens.

Izawati AM, Masani MY, Ismanizan I, Parveez GK - Front Plant Sci (2015)

Schematic diagram of DOGR1 expression cassette (line indicates the size of 1.5 kb) used as probe for Southern blot analysis. RB, right border; LB, left border; NOS, nopaline synthase gene terminator; DOGR1, gene codes for 2-deoxyglucose-6-phosphate phosphatase; CaMV35S, cauliflower mosaic virus 35S promoter. Arrows indicates the position of primers used for PCR analysis to amplify 741 bp.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585222&req=5

Figure 1: Schematic diagram of DOGR1 expression cassette (line indicates the size of 1.5 kb) used as probe for Southern blot analysis. RB, right border; LB, left border; NOS, nopaline synthase gene terminator; DOGR1, gene codes for 2-deoxyglucose-6-phosphate phosphatase; CaMV35S, cauliflower mosaic virus 35S promoter. Arrows indicates the position of primers used for PCR analysis to amplify 741 bp.
Mentions: The construction of pBIDOG plasmid was performed by replacing the DNA fragment of CaMV35S-GUS-Nos of pBI35SGUS with the DNA fragment of CaMV35S-DOGR1-Nos of pBINARDOG (SunGene, Germany). pBI121 plasmid was digested with PmeI and ClaI to remove the DNA fragment of NosPro-nptII-Nos, rendered blunt and religated to generate plasmid pBI35SGUS. The CaMV35S-DOGR1-Nos fragment was released from pBINARDOG by HindIII and EcoRI digestion, and cloned into pBI35SGUS releasing the DNA fragment of CaMV35S-GUS-Nos at similar sites to generate pBIDOG (Figure 1). pBIDOG was transformed into E. coli and later mobilized into Agrobacterium tumefaciens strain LBA4404 via electroporation and used as a vector for oil palm transformation.

Bottom Line: The plantlets were later transferred into soil and grown in a biosafety screenhouse.PCR and subsequently Southern blot analyses were carried out to confirm the integration of the transgene in the plantlets.This result demonstrates the potential of using combination of DOG(R)1 gene and 2-DOG for regenerating transgenic oil palm.

View Article: PubMed Central - PubMed

Affiliation: Advanced Biotechnology and Breeding Centre, Malaysian Palm Oil Board Selangor, Malaysia.

ABSTRACT
DOG(R)1, which encodes 2-deoxyglucose-6-phosphate phosphatase, has been used as a selectable marker gene to produce transgenic plants. In this study, a transformation vector, pBIDOG, which contains the DOG(R)1 gene, was transformed into oil palm embryogenic calli (EC) mediated by Agrobacterium tumefaciens strain LBA4404. Transformed EC were exposed to 400 mg l(-1) 2-deoxyglucose (2-DOG) as the selection agent. 2-DOG resistant tissues were regenerated into whole plantlets on various regeneration media containing the same concentration of 2-DOG. The plantlets were later transferred into soil and grown in a biosafety screenhouse. PCR and subsequently Southern blot analyses were carried out to confirm the integration of the transgene in the plantlets. A transformation efficiency of about 1.0% was obtained using DOG(R)1 gene into the genome of oil palm. This result demonstrates the potential of using combination of DOG(R)1 gene and 2-DOG for regenerating transgenic oil palm.

No MeSH data available.


Related in: MedlinePlus