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Identification and characterization of microRNAs from citrus expressed sequence tags

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Abstract

MicroRNAs (miRNAs) are a group of single-stranded noncoding RNAs with general size of 21–24 nucleotides, which negatively regulate gene expression posttranscriptionally by repressing gene translation or degrading targeted mRNAs. Despite the important functions of miRNAs in plants, little is known about miRNAs in citrus. Here we present a study of bioinformatics identification of microRNA precursors in citrus by comparing known plant miRNAs against 560,271 citrus ESTs. Seventy-eight ESTs from ten citrus species and three hybrids were identified as putative miRNA precursors, encoding 51 unique miRNA sequences, representing 28 miRNA families. Blastn search of the putative miRNAs against citrus unigenes and ESTs was performed to identify the target genes. As a result, 36 unigenes and 77 ESTs were identified as putative targets of 25 citrus miRNA families. The putative targets are mainly transcription factors and play important roles in development, metabolism, and stress resistance. To validate the predicted miRNAs, qRT-PCR was applied to detect the tissue-specific expression of nine putative miRNAs in Citrus sinensis cv. Valencia. The cleavage sites on five mRNAs targeted by three miRNA families were mapped by 5′RACE. This study provided information on citrus miRNA precursors, mature miRNAs, and miRNA targets and is helpful for future research of miRNA function in citrus.

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Abbreviations

EST:

expressed sequence tag

miRNA:

microRNA

miRNA*:

passenger strand on precursor

Pre-miRNA:

microRNA precursor

MFE:

negative minimal free energies

MFEi:

minimal free energy index

qRT-PCR:

quantitative reverse transcription-polymerase chain reation

5′RACE:

5′ rapid amplification of cDNA ends

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Acknowledgements

This research was financially supported by the National Natural Science Foundation (No. 30921002), the Ministry of Agriculture of China (No. 200903044-3), the Fundamental Research Funds for the Central Universities (No. 2009PY022) and the Hubei provincial Natural Science Foundation (No. 2008CDA069). The authors thank Dr. Han-Hui Kuang (Huazhong Agric. Univ.) for his critical reading the manuscript.

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  1. Key Laboratory of Horticultural Plant Biology (Ministry of Education), National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, 430070, China

    Xiao-Meng Wu, Mei-Ya Liu, Qiang Xu & Wen-Wu Guo

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  1. Xiao-Meng Wu
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  2. Mei-Ya Liu
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  3. Qiang Xu
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  4. Wen-Wu Guo
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Correspondence to Wen-Wu Guo.

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Communicated by F. Gmitter

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Wu, XM., Liu, MY., Xu, Q. et al. Identification and characterization of microRNAs from citrus expressed sequence tags. Tree Genetics & Genomes 7, 117–133 (2011). https://doi.org/10.1007/s11295-010-0319-5

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