In vitro transcription of chloroplast protein genes
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This chapter discusses the methods that are useful for the in vitro analysis of plastid gene expression. Many plastid genes have multiple transcripts in vivo that may be the result of transcription initiation and subsequent RNA processing events. Heterologous chloroplast transcription extracts may be useful for the analysis of plastid RNA processing. To characterize the potential regulatory nature of the plastid RNA and other plastid DNA sequences, the use of a homologous plastid in vitro transcription system is required. The criteria for such a transcription system are ease of preparation, accuracy of transcription initiation, and stability of enzyme activity. The optimal time and temperature of the in vitro transcription reaction varies with the gene transcribed and the extract employed. Part of this variance may be due to specific RNA processing and nonspecific degradation of in vitro transcripts by endogenous ribonucleases in the chloroplast extracts. © 1986, Elsevier Inc.
author list (cited authors)
Orozco, E. M., Mullet, J. E., Hanley-Bowdoin, L., & Chua, N.