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Characterization and expression analysis of waxy alleles in barley accessions

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Abstract

Granule Bound Starch Synthase I (GBSS I) encoded by the waxy gene plays an important role in accumulating amylose during the development of starch granules in barley. In this study, we isolated and characterized waxy alleles of three waxy (GSHO 908, GSHO 1828 and NA 40) and two non-waxy barley accessions (PI 483237 and CIho 15773), estimated the expression patterns of waxy genes via Real-time quantitative PCR (RT-qPCR), investigated promoter activity by analyzing promoter-GUS expression, and examined possible effects of waxy alleles on starch granule morphology in barley accessions by scanning electron microscopy (SEM). A 193-bp insertion in intron 1, a 15-bp insertion in the coding region, and some single nucleotide polymorphic sites were detected in the waxy barley accessions. In addition, a 397-bp deletion containing the TATA box, transcription starting point, exon 1 and partial intron 1 were also identified in the waxy barley accessions. RT-qPCR analysis showed that waxy accessions had lower waxy expression levels than those of non-waxy accessions. Transient expression assays showed that GUS activity driven by the 1,029-bp promoter of the non-waxy accessions was stronger than that driven by the 822-bp promoter of the waxy accessions. SEM revealed no apparent differences of starch granule morphology between waxy and non-waxy accessions. Our results showed that the 397-bp deletion identified in the waxy barley accessions is likely responsible for the reduction of waxy transcript, leading to lower concentrations of GBSS I protein thus lower amylose content.

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (31230053, 31000167 and 31171556) and China Transgenic Research Program (2011ZX08002-001,004 and 005) the National Basic Research Program of China (973 Program 2010CB134400).

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Authors and Affiliations

  1. Triticeae Research Institute, Sichuan Agricultural University, Chengdu, 611130, Sichuan, China

    Jian Ma, Qian-Tao Jiang, Quan-Zhi Zhao, Shan Zhao, Xiu-Jin Lan, Shou-Fen Dai & Yu-Ming Wei

  2. Lethbridge Research Centre, Agriculture and Agri-Food Canada, Lethbridge, T1J4B1, Canada

    Zhen-Xiang Lu

  3. CSIRO Plant Industry, 306 Carmody Road, St. Lucia, QLD, 4067, Australia

    Chunji Liu

  4. Key Laboratory of Southwestern Crop Germplasm Utilization, Ministry of Agriculture, Sichuan Agricultural University, Ya’an, 625014, Sichuan, China

    You-Liang Zheng

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  1. Jian Ma
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Correspondence to You-Liang Zheng.

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Jian Ma and Qian-Tao Jiang contributed equally to this paper.

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Ma, J., Jiang, QT., Zhao, QZ. et al. Characterization and expression analysis of waxy alleles in barley accessions. Genetica 141, 227–238 (2013). https://doi.org/10.1007/s10709-013-9721-x

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