Genetic transformation and molecular analysis of polyhydroxybutyrate biosynthetic gene expression in oil palm (Elaeis guineensis Jacq. var Tenera) tissues

Ismanizan Ismail, Nor Fakhrana Iskandar, Gor Mian Chee, Ruslan Abdullah

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Bioplastics are an alternative substitute for petrochemical synthetic plastics. Polyhydroxybutyrate (PHB) genes are involved in bioplastic synthesis. In this study, bioplastic synthesis genes were incorporated into the genome of oil palm because this plant has a high concentration of the PHB precursor acetyl-CoA. Immature embryos (IEs) of Elaeis guineensis var Tenera were infected with Agrobacterium tumefaciens LBA4404 that contained the binary vector pJLPHB3, which encoded the phb genes, β-ketothiolase (bktB), acetoacetyl-CoA reductase (phaB) and PHA synthase (phaC) flanked by a modified CaMV35S promoter, a plastid targeting sequence and the nos terminator. GUS assay revealed that about 78-100% transient transformation frequency was obtained for calluses and 55-65% for plantlets 1 month after transformation. However, GUS assays of leaf tissue from 12-month-old plantlets showed that only 10-33% transformation frequency was obtained. The presence of the phb genes in GUS positive plantlets was confirmed using PCR and multiplex PCR analyses. Southern blot analyses verified that the phb genes were integrated in transformed leaves and calluses using the phaB probe (0.805 kb) and phaC probe (1.730 kb). Quantitative transgene expression comparison in the transformed tissues measured using real-time PCR showed that the expression levels of the phaB and phaC transgenes were 6.06- and 6.02-fold higher compared to the non-transformed oil palm.

Original languageEnglish
Pages (from-to)18-27
Number of pages10
JournalPlant OMICS
Volume3
Issue number1
Publication statusPublished - Jan 2010

Fingerprint

polyhydroxybutyrate
Elaeis guineensis
genetic transformation
gene expression
plantlets
genes
transgenes
callus
petrochemicals
synthesis
assays
Agrobacterium radiobacter
embryo (plant)
Southern blotting
plastids
leaves
quantitative polymerase chain reaction
plastics
immatures
promoter regions

Keywords

  • Agrobacterium tumefaciens
  • Binary vector
  • Elaeis guineensis
  • Genetic transformation
  • Immature embryo
  • Polyhydroxybutyrate

ASJC Scopus subject areas

  • Agronomy and Crop Science
  • Plant Science

Cite this

Genetic transformation and molecular analysis of polyhydroxybutyrate biosynthetic gene expression in oil palm (Elaeis guineensis Jacq. var Tenera) tissues. / Ismail, Ismanizan; Iskandar, Nor Fakhrana; Chee, Gor Mian; Abdullah, Ruslan.

In: Plant OMICS, Vol. 3, No. 1, 01.2010, p. 18-27.

Research output: Contribution to journalArticle

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abstract = "Bioplastics are an alternative substitute for petrochemical synthetic plastics. Polyhydroxybutyrate (PHB) genes are involved in bioplastic synthesis. In this study, bioplastic synthesis genes were incorporated into the genome of oil palm because this plant has a high concentration of the PHB precursor acetyl-CoA. Immature embryos (IEs) of Elaeis guineensis var Tenera were infected with Agrobacterium tumefaciens LBA4404 that contained the binary vector pJLPHB3, which encoded the phb genes, β-ketothiolase (bktB), acetoacetyl-CoA reductase (phaB) and PHA synthase (phaC) flanked by a modified CaMV35S promoter, a plastid targeting sequence and the nos terminator. GUS assay revealed that about 78-100{\%} transient transformation frequency was obtained for calluses and 55-65{\%} for plantlets 1 month after transformation. However, GUS assays of leaf tissue from 12-month-old plantlets showed that only 10-33{\%} transformation frequency was obtained. The presence of the phb genes in GUS positive plantlets was confirmed using PCR and multiplex PCR analyses. Southern blot analyses verified that the phb genes were integrated in transformed leaves and calluses using the phaB probe (0.805 kb) and phaC probe (1.730 kb). Quantitative transgene expression comparison in the transformed tissues measured using real-time PCR showed that the expression levels of the phaB and phaC transgenes were 6.06- and 6.02-fold higher compared to the non-transformed oil palm.",
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