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    Home > Biochemistry News > Plant Extracts News > Cloning New Small RNA Sequences

    Cloning New Small RNA Sequences

    • Last Update: 2020-11-06
    • Source: Internet
    • Author: User
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    Small RNAs are key molecules in RNA silencing pathways that exert sequence-specific regulation of gene expression and chromatin modifications in many eukaryotes. In plants, endogenous small RNAs, including microRNAs (miRNAs) and
    trans
    -acting small interfering RNAs (tasiRNAs) play an important role in biological processes such as development and stress responses. In addition, viral genome-derived siRNAs are produced during viral infection, and they exhibit anti-viral defense by an RNA silencing pathway. These endogenous and exogenous small RNAs are mainly 21-24 nucleotides in length. Here, we describe a method to identify small RNA sequences from plant tissues. Small RNAs are purified by column fractionation and gel excision from total RNAs. These small RNAs are ligated at both termini to
    DNA
    /RNA chimeric adapters and reverse-transcribed to produce
    cDNA
    s. By the following
    PCR
    amplification,
    Ban
    I restriction sites are added to cDNAs, which enables directional concatamerization. Concatamerized-fragments are cloned and sequenced. This method could be applied to identify small RNA sequences from many sources, e.g., mutant plants, plants in various stress environments, and virus-infected plants.
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