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Determination of the Half-Life of Chloroplast Transcripts in Tobacco Leaves

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Chloroplast Biotechnology

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1132))

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Abstract

The amounts of specific transcripts that accumulate in chloroplasts are determined by the rates of synthesis and degradation of the transcripts. The 3′ untranslated region of transcripts is a major determinant of the stability of transcripts in chloroplasts. The half-lives of specific transcripts can be determined by northern blot analysis of a time course of transcripts in detached tobacco leaves incubated with actinomycin D, a potent transcription inhibitor. This analysis may be applied to transcripts of endogenous genes or of transgenes introduced into the chloroplast genome in transplastomic plants. Sequence determinants of transcript stability can be identified by analysis of transplastomic plants containing constructs of the green fluorescent protein (gfp) reporter gene fused to the sequences of interest.

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Acknowledgments

We thank Sue Aspinall for help in writing this article and for laboratory assistance during the development of the methods described. S.T. was supported by a scholarship from the Thai Government during the development of the methods described.

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Authors and Affiliations

  1. National Center for Genetic Engineering and Biotechnology, Klong Luang, Pathumthani, Thailand

    Sithichoke Tangphatsornruang

  2. Department of Plant Sciences, University of Cambridge, Cambridge, UK

    John C. Gray

Authors
  1. Sithichoke Tangphatsornruang
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  2. John C. Gray
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Editor information

Editors and Affiliations

  1. Rutgers University, Waksman Inst.of Microbiology, Piscataway, New Jersey, USA

    Pal Maliga

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Tangphatsornruang, S., Gray, J.C. (2014). Determination of the Half-Life of Chloroplast Transcripts in Tobacco Leaves. In: Maliga, P. (eds) Chloroplast Biotechnology. Methods in Molecular Biology, vol 1132. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-995-6_13

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  • DOI: https://doi.org/10.1007/978-1-62703-995-6_13

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-994-9

  • Online ISBN: 978-1-62703-995-6

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