Epigenetic transcriptional silencing and 5-azacytidine-mediated reactivation of a complex transgene in rice

Siva P. Kumpatla, Weimin Teng, Wallace G Buchholz, Timothy C. Hall

Research output: Contribution to journalArticle

103 Citations (Scopus)

Abstract

Despite a growing number of reports indicating non-Mendelian inheritance of transgene expression in monocots, no detailed description of the structure and stability of the transgene exists for transformants generated by direct DNA-transfer techniques, making the cause for these observations difficult to determine. In this paper we describe the complex organization of Btt cryIIIA and bar transgenes in rice (Oryza sativa L.) that displayed aberrant segregation in R1 progeny. Silencing rather than rearrangement of the bar gene was implicated because the herbicide-sensitive R1 plants had a DNA hybridization profile identical to that of the resistant R0 parent and R1 siblings. Genomic DNA analysis revealed substantial methylation of the Ubi1/bar sequences in silenced plants and, to a lesser degree, in herbicide-resistant plants, suggesting that the transgene locus was potentiated for silencing. Nuclease protection and nuclear run-on assays confirmed that silencing was due to transcriptional inactivation. Treatment of R2 progeny of silenced plants with 5-azacytidine resulted in demethylation of the Ubi1 promoter and reactivation of bar gene expression, demonstrating a functional relationship for methylation in gene silencing. These findings indicate that methylation-based silencing may be frequent in cereals transformed by direct DNA protocols that insert multiple, often rearranged sequences.

Original languageEnglish (US)
Pages (from-to)361-373
Number of pages13
JournalPlant Physiology
Volume115
Issue number2
DOIs
StatePublished - Jan 1 1997

Fingerprint

Azacitidine
Transgenes
Epigenomics
epigenetics
transgenes
methylation
Methylation
rice
DNA
Herbicides
herbicides
transfer DNA
Gene Rearrangement
nucleases
Gene Silencing
gene silencing
Liliopsida
nucleic acid hybridization
inheritance (genetics)
Oryza sativa

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Cite this

Epigenetic transcriptional silencing and 5-azacytidine-mediated reactivation of a complex transgene in rice. / Kumpatla, Siva P.; Teng, Weimin; Buchholz, Wallace G; Hall, Timothy C.

In: Plant Physiology, Vol. 115, No. 2, 01.01.1997, p. 361-373.

Research output: Contribution to journalArticle

Kumpatla, Siva P. ; Teng, Weimin ; Buchholz, Wallace G ; Hall, Timothy C. / Epigenetic transcriptional silencing and 5-azacytidine-mediated reactivation of a complex transgene in rice. In: Plant Physiology. 1997 ; Vol. 115, No. 2. pp. 361-373.
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