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Journal of Integrative Agriculture  2017, Vol. 16 Issue (03): 572-579    DOI: 10.1016/S2095-3119(16)61460-0
Crop Genetics · Breeding · Germplasm Resources Advanced Online Publication | Current Issue | Archive | Adv Search |
Evaluation of parameters affecting Agrobacterium-mediated transient expression in citrus
LI Fang1*, DAI Su-ming1*, DENG Zi-niu1, LI Da-zhi1, LONG Gui-you1, LI Na1, LI Yi1, 2, Alexandra Gentile1, 3

1 National Center for Citrus Improvement Changsha/Horticulture and Landscape College, Hunan Agricultural University, Changsha 410128, P.R.China

2 Department of Plant Science, University of Connecticut, Connecticut 06269, USA

3 Department of Agriculture and Food Science, University of Catania, Catania 95123, Italy

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Abstract  Agrobacterium-mediated transient expression assays are a convenient alternative to stable expression because they are simple, easy to perform, and achieve gene expression rapidly.  This study investigated the factors affecting transient gene expression efficiency in citrus by observing the cryo-sectioning of leaf samples under a laser confocal microscope.  These factors included the composition of the infiltration buffer, the Agrobacterium cell density, the leaf development stage, the incubation temperature, and plant genotype.  The highest transient expression level of yellow fluorescent protein (YFP) was detected in Mexican lime (Citrus aurantifolia) on the third day after the intermediate-aged leaves were infiltrated with the improved infiltration buffer 1 (15 mmol L-1 2-(N-morpholino) ethanesulfonic acid, 10 mmol L-1 MgCl2, and 200 μmol L-1 acetosyringone), which had an optical density of 0.8 and was incubated at 22°C.  Additionally, this transient expression assay was applied to other citrus genotypes.  Of note, trifoliate orange (Poncirus trifoliata) and kumquat (Fortunella obovate) had higher expression efficiency than other six genotypes of the Citrus genus.  Our study provides research basis for the selection of optimization strategies in transient gene expression and improves the method for available genome investigation in citrus.
Keywords:  citrus      Agrobacterium      transient expression efficiency      yellow fluorescent protein (YFP)  
Received: 05 March 2016   Accepted:
Fund: 

This study was financed by the National Natural Science Foundation of China (30900972, 31572111), the Special Found for Agro-scientific Research in the Public Interest, China (201203076-06) and the Graduate Innovative Projects of Hunan Province, China (CX2013B290).

Corresponding Authors:  DAI Su-ming, Tel: +86-731-84638214, Fax: +86-731-84635302, E-mail: dsm531@126.com* These authors contributed    
About author:  LI Fang, E-mail: lifang200709@126.com

Cite this article: 

LI Fang, DAI Su-ming, DENG Zi-niu, LI Da-zhi, LONG Gui-you, LI Na, LI Yi, Alexandra Gentile. 2017. Evaluation of parameters affecting Agrobacterium-mediated transient expression in citrus. Journal of Integrative Agriculture, 16(03): 572-579.

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