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Two Highly Homologous Promoters of a Squash Aspartic Protease Inhibitor (SQAPI) Multigene Family Exhibit Differential Expression in Transgenic Tobacco Phloem and Trichome Cells

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Abstract

Two variants of the promoter of the squash aspartic acid protease inhibitor multigene family were isolated from Cucurbita maxima cv. ‘Supermarket’ Hybrid genomic DNA. The isolated promoters, possibly not full length, comprised a 5′-untranslated region (UTR) of 202–208 bp, contained a 63-bp upstream open reading frame (uORF) and the immediate upstream sequences of 441–445 bp. The two promoters contained several small deletions relative to each other and 22 single base differences but exhibit overall 92.5% homology over 654 bp. When the promoters were fused to a β-glucuronidase reporter gene and expressed in tobacco, one variant was highly expressed in the companion cells of the inner and outer phloem of leaves and at lower levels in other organs. The other variant was expressed at high levels in the long glandular trichomes of the leaf. Deletion analysis identified a region of ~280 bp immediately upstream of the 5′-UTR containing the TATA box that was responsible for phloem specific expression and a further region of ~180 bp that enhanced expression in one promoter and conferred trichome expression in the other. Removal of the 5′-UTR, including the uORF, inactivated the phloem promoter.

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Abbreviations

SQAPI:

Squash aspartic protease inhibitor

TSS:

Transcription start site

uORF:

Upstream open reading frame

5′-UTR:

5′-untranslated region

GUS:

β-Glucuronidase

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Acknowledgments

We would like to thank Douglas Hopcroft and Raymond Bennett for assistance with microscopy. The research was made possible by a grant from the New Zealand Public Good Research Fund (FfRST Contract CO6X0207).

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  1. The New Zealand Institute of Plant and Food Research Ltd, Private Bag 11030, Palmerston North, 4442, New Zealand

    Ananda Anandan, Laurence N. Gatehouse, Richelle K. Marshall, Colleen Murray & John T. Christeller

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  1. Ananda Anandan
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  2. Laurence N. Gatehouse
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Correspondence to John T. Christeller.

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Anandan, A., Gatehouse, L.N., Marshall, R.K. et al. Two Highly Homologous Promoters of a Squash Aspartic Protease Inhibitor (SQAPI) Multigene Family Exhibit Differential Expression in Transgenic Tobacco Phloem and Trichome Cells. Plant Mol Biol Rep 27, 355–364 (2009). https://doi.org/10.1007/s11105-009-0096-1

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