Expression of a synthetic E. coli heat-labile enterotoxin B sub-unit (LT-B) in maize

Rachel Chikwamba, Jennifer McMurray, Huixia Shou, Bronwyn Frame, Sue Ellen Pegg, Paul Scott, Hugh Mason, Kan Wang

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43 Scopus citations

Abstract

We have produced the B subunit of the enterotoxigenic Escherichia coli (ETEC) heat-labile enterotoxin (LT-B) in transgenic maize seed. LT-B is a model antigen that induces a strong immune response upon oral administration and enhances immune responses to conjugated and co-administered antigens. Using a synthetic LT-B gene with optimized codon sequence, we examined the role of promoters and the SEKDEL endoplasmic reticulum retention motif in LT-B accumulation in callus and in kernels. Two promoters, the constitutive CaMV 35S promoter and the maize 27 kDa gamma zein promoter, which directs endosperm-specific gene expression in maize kernels, regulated LT-B expression. Ganglioside-dependent ELISA analysis showed that using the constitutive promoter, maximum LT-B level detected in callus was 0.04% LT-B in total aqueous-extractable protein (TAEP) and 0.01% in R1 kernels of transgenic plants. Using the gamma zein promoter, LT-B accumulation reached 0.07% in R1 kernels. The SEKDEL resulted in increased LT-B levels when combined with the gamma zein promoter. We monitored LT-B levels under greenhouse and field conditions over three generations. Significant variability in gene expression was observed between transgenic events, and between plants within the same event. A maximum of 0.3% LT-B in TAEP was measured in R3 seed of a transgenic line carrying CaMV 35S promoter/LT-B construct. In R3 seed of a transgenic line carrying the gamma zein promoter/LT-B construct, up to 3.7% LT-B in TAEP could be detected. We concluded that maize seed can be used as a production system for functional antigens.

Original languageEnglish (US)
Pages (from-to)253-265
Number of pages13
JournalMolecular Breeding
Volume10
Issue number4
DOIs
StatePublished - Nov 25 2002

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Keywords

  • Functional antigen
  • Gene expression
  • LT-B
  • Zea mays

ASJC Scopus subject areas

  • Biotechnology
  • Molecular Biology
  • Agronomy and Crop Science
  • Genetics
  • Plant Science

Cite this

Chikwamba, R., McMurray, J., Shou, H., Frame, B., Pegg, S. E., Scott, P., Mason, H., & Wang, K. (2002). Expression of a synthetic E. coli heat-labile enterotoxin B sub-unit (LT-B) in maize. Molecular Breeding, 10(4), 253-265. https://doi.org/10.1023/A:1020509915672