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Identification of soybean microRNAs and their targets

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Abstract

The microRNAs (miRNAs) are a newly identified class of small non-protein-coding regulatory RNA. Using comparative genomics, we identified 69 miRNAs belonging to 33 families in the domesticated soybean (Glycine max) as well as five miRNAs in the soybean wild species Glycine soja and Glycine clandestine. TaqMan® MicroRNA Assay analyses demonstrated that these miRNAs were differentially expressed in soybean tissues, with certain classes expressed preferentially in both a spatiotemporal and a tissue-specific manner. Detailed sequence analyses revealed that soybean pre-miRNAs vary in length from 44 to 259 nt with an average of 106 ± 45 nt, harbor mature miRNAs that differ in their physical location within the pre-miRNAs, and encode more than a single mature miRNA. Comparative sequence analyses of soybean miRNA sequences showed that uracil is the dominant base in the first position at the 5′ end of the mature miRNAs while cytosine is dominant at the 19th position, which is indicative that these two bases may have an important functional role in miRNA biogenesis and/or miRNA-mediated gene regulation. Soybeans were unique among plants in the frequency of occurrence of miRNA clusters. For the first time, antisense miRNAs were identified in plants. The five antisense miRNAs and their sense partners from soybean belonged to three miRNA families (miR-157, miR-162 and miR-396). Antisense miRNAs were also identified in soybean wild species. Mature antisense miRNA products appeared to have 1–3 nucleotide changes compared to their sense partners, which suggests that both strands of a miRNA gene can produce functional mature miRNAs and that antisense transcripts may differ functionally from their sense partners. Based on previously established in silico methods, we predicted 152 miRNA-targeted mRNAs, which included a large percentage of mRNAs that encode transcription factors that regulate plant growth and development as well as a lesser percentage of mRNAs that encode environmental signal transduction proteins and central metabolic processes.

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Abbreviations

3′ UTR:

3′ Untranslated region

AMFE:

Adjusted MFE

ap2 :

Apetal 2

ARFs:

Auxin response factors

CBF:

CCAAT-binding transcription factor

EST:

Expressed sequence tag

GSS:

Genome survey sequence

GIF1:

GRF1-interacting factor 1

HSP:

Heat shock protein

miRNA:

MicroRNA

MFEI:

Minimal folding free energy index

pre-miRNAs:

miRNA precusor

NCBI:

National Center for Biotechnology Information

NAP:

Nucleosome assembly protein

PCR:

Polymerase chain reaction

qRT-PCR:

Quantitative real time PCR

RISC:

RNA-induced silencing complex

SBP:

Squamosa-promoter binding protein-like protein

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Acknowledgments

This work was supported by East Carolina University New Faculty Research Startup Funds Program (BZ), Western Illinois University New Faculty Research Startup Funds Program (XP), and a Science and Engineering Grant from DuPont (BZ). We would like to thank Dr. Joseph Burton of ARS, USDA for kindly providing soybean seeds. We also appreciate Dr. Brian McGonigle of DuPont for his support and encouragement.

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Authors and Affiliations

  1. Department of Biology, East Carolina University, Greenville, NC, 27858, USA

    Baohong Zhang & Edmund J. Stellwag

  2. Department of Chemistry, Western Illinois University, Macomb, IL, 61455, USA

    Xiaoping Pan

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  1. Baohong Zhang
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  2. Xiaoping Pan
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Correspondence to Baohong Zhang.

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425_2008_818_MOESM1_ESM.doc

MOESM1 Schematic representation of the miRNA gene search procedure for identifying soybeanhomologs based on established Arabidopsis miRNAs (DOC 121 kb)

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Zhang, B., Pan, X. & Stellwag, E.J. Identification of soybean microRNAs and their targets. Planta 229, 161–182 (2008). https://doi.org/10.1007/s00425-008-0818-x

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