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High-efficiency RNA cloning enables accurate quantification of miRNA expression by deep sequencing.

ABSTRACT: Small RNA cloning and sequencing is uniquely positioned as a genome-wide approach to quantify miRNAs with single-nucleotide resolution. However, significant biases introduced by RNA ligation in current protocols lead to inaccurate miRNA quantification by 1000-fold. Here we report an RNA cloning method that achieves over 95% efficiency for both 5? and 3? ligations. It achieves accurate quantification of synthetic miRNAs with less than two-fold deviation from the anticipated value and over a dynamic range of four orders of magnitude. Taken together, this high-efficiency RNA cloning method permits accurate genome-wide miRNA profiling from total RNAs.

SUBMITTER: Zhang Z

PROVIDER: S-EPMC3983620 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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High-efficiency RNA cloning enables accurate quantification of miRNA expression by deep sequencing.

Zhang Zhaojie Z Lee Jerome E JE Riemondy Kent K Anderson Emily M EM Yi Rui R

Genome biology 20130101 10

Small RNA cloning and sequencing is uniquely positioned as a genome-wide approach to quantify miRNAs with single-nucleotide resolution. However, significant biases introduced by RNA ligation in current protocols lead to inaccurate miRNA quantification by 1000-fold. Here we report an RNA cloning method that achieves over 95% efficiency for both 5′ and 3′ ligations. It achieves accurate quantification of synthetic miRNAs with less than two-fold deviation from the anticipated value and over a dynam ...[more]

PMID: 24098942

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High-efficiency RNA cloning enables quantitative measurement of miRNA expression by deep sequencing

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