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Open Access Highly Accessed Research article

Concerted suppression of all starch branching enzyme genes in barley produces amylose-only starch granules

Massimiliano Carciofi1, Andreas Blennow2*, Susanne L Jensen23, Shahnoor S Shaik2, Anette Henriksen4, Alain Buléon5, Preben B Holm1 and Kim H Hebelstrup1*

Author affiliations

1 Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark

2 Department of Plant Biology and Biotechnology, VKR Research Centre for Pro-Active Plants, Faculty of Life Sciences, University of Copenhagen, Frederiksberg, Denmark

3 KMC, Herningvej 60, Brande, 7330, Denmark

4 The Protein Chemistry Group, Carlsberg Laboratory, Copenhagen, Denmark

5 UR1268 Biopolymeres Interactions Assemblages, INRA, Nantes, F-44300, France

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Citation and License

BMC Plant Biology 2012, 12:223  doi:10.1186/1471-2229-12-223

Published: 21 November 2012

Abstract

Background

Starch is stored in higher plants as granules composed of semi-crystalline amylopectin and amorphous amylose. Starch granules provide energy for the plant during dark periods and for germination of seeds and tubers. Dietary starch is also a highly glycemic carbohydrate being degraded to glucose and rapidly absorbed in the small intestine. But a portion of dietary starch, termed “resistant starch” (RS) escapes digestion and reaches the large intestine, where it is fermented by colonic bacteria producing short chain fatty acids (SCFA) which are linked to several health benefits. The RS is preferentially derived from amylose, which can be increased by suppressing amylopectin synthesis by silencing of starch branching enzymes (SBEs). However all the previous works attempting the production of high RS crops resulted in only partly increased amylose-content and/or significant yield loss.

Results

In this study we invented a new method for silencing of multiple genes. Using a chimeric RNAi hairpin we simultaneously suppressed all genes coding for starch branching enzymes (SBE I, SBE IIa, SBE IIb) in barley (Hordeum vulgare L.), resulting in production of amylose-only starch granules in the endosperm. This trait was segregating 3:1. Amylose-only starch granules were irregularly shaped and showed peculiar thermal properties and crystallinity. Transgenic lines retained high-yield possibly due to a pleiotropic upregualtion of other starch biosynthetic genes compensating the SBEs loss. For gelatinized starch, a very high content of RS (65 %) was observed, which is 2.2-fold higher than control (29%). The amylose-only grains germinated with same frequency as control grains. However, initial growth was delayed in young plants.

Conclusions

This is the first time that pure amylose has been generated with high yield in a living organism. This was achieved by a new method of simultaneous suppression of the entire complement of genes encoding starch branching enzymes. We demonstrate that amylopectin is not essential for starch granule crystallinity and integrity. However the slower initial growth of shoots from amylose-only grains may be due to an important physiological role played by amylopectin ordered crystallinity for rapid starch remobilization explaining the broad conservation in the plant kingdom of the amylopectin structure.

Keywords:
Amylose-only starch; Resistant starch; RNA interference; Starch branching enzymes; Starch bioengineering; Starch granules; Starch crystallinity; Barley