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The sucrose synthase-1 promoter from Citrus sinensis directs expression of the β-glucuronidase reporter gene in phloem tissue and in response to wounding in transgenic plants

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Abstract

Interest in phloem-specific promoters for the engineering of transgenic plants has been increasing in recent years. In this study we isolated two similar, but distinct, alleles of the Citrus sinensis sucrose synthase-1 promoter (CsSUS1p) and inserted them upstream of the β-glucuronidase (GUS) gene to test their ability to drive expression in the phloem of transgenic Arabidopsis thaliana and Nicotiana tabacum. Although both promoter variants were capable of conferring localized GUS expression in the phloem, the CsSUS1p-2 allele also generated a significant level of expression in non-target tissues. Unexpectedly, GUS expression was also instigated in a minority of CsSUS1p::GUS lines in response to wounding in the leaves of transgenic Arabidopsis. Deletion analysis of the CsSUS1p suggested that a fragment comprising nucleotides −410 to −268 relative to the translational start site contained elements required for phloem-specific expression while nucleotides −268 to −103 contained elements necessary for wound-specific expression. Interestingly, the main difference between the two CsSUS1p alleles was the presence of a 94-bp insertion in allele 2. Fusion of this indel to a minimal promoter and GUS reporter gene indicated that it contained stamen and carpel-specific enhancer elements. This finding of highly specific and separable regulatory units within the CsSUS1p suggests that this promoter may have a potential application in the generation of constructs for the use in the development of transgenic plants resistant to a wide variety of target pests.

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Abbreviations

BA:

Benzyl adenine

CaMV:

Cauliflower mosaic virus

CsSUS1p :

SUS1 upstream region from Citrus sinensis

GUS:

β-Glucuronidase

MS:

Murashige and Skoog

MU:

Methylumbelliferone

PCR:

Polymerase chain reaction

5′ RACE:

5′ Rapid amplification of cDNA ends

5′ RAGE:

5′ Rapid amplification of genomic DNA ends

RT-PCR:

Reverse transcription polymerase chain reaction

SUS:

Sucrose synthase

UDP:

Uridine diphosphate

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Acknowledgments

This study was supported in part by state, federal, and institutional funds appropriated to the New York State Agricultural Experiment Station, Cornell University, and by the State of Florida Department of Citrus and the Citrus Research and Development Foundation, Inc. project NAS ID number 38 (Cornell OSP 57390 and 63013). We also thank Sara Villani for her invaluable technical assistance.

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  1. Department of Plant Pathology and Plant–Microbe Biology, New York State Agricultural Experiment Station, Cornell University, Geneva, NY, 14456, USA

    Stacy D. Singer, Jean-Michel Hily & Kerik D. Cox

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  1. Stacy D. Singer
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Correspondence to Kerik D. Cox.

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Singer, S.D., Hily, JM. & Cox, K.D. The sucrose synthase-1 promoter from Citrus sinensis directs expression of the β-glucuronidase reporter gene in phloem tissue and in response to wounding in transgenic plants. Planta 234, 623–637 (2011). https://doi.org/10.1007/s00425-011-1432-x

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