A functional assay for microRNA target identification and validation

Joop Gaken; Azim M Mohamedali; Jie Jiang; Farooq Malik; Doris Stangl; Alexander E Smith; Constantinos Chronis; Austin G Kulasekararaj; N Shaun B Thomas; Farzin Farzaneh; Mahvash Tavassoli; Ghulam J Mufti

doi:10.1093/nar/gks145

A functional assay for microRNA target identification and validation

Joop Gaken, Azim M Mohamedali, Jie Jiang, Farooq Malik, Doris Stangl, Alexander E Smith, Constantinos Chronis, Austin G Kulasekararaj, N Shaun B Thomas, Farzin Farzaneh, Mahvash Tavassoli, Ghulam J Mufti

Centre for Host Microbiome Interactions

Research output: Contribution to journal › Article › peer-review

Abstract

MicroRNAs (miRNA) are a class of small RNA molecules that regulate numerous critical cellular processes and bind to partially complementary sequences resulting in down-regulation of their target genes. Due to the incomplete homology of the miRNA to its target site identification of miRNA target genes is difficult and currently based on computational algorithms predicting large numbers of potential targets for a given miRNA. To enable the identification of biologically relevant miRNA targets, we describe a novel functional assay based on a 3'-UTR-enriched library and a positive/negative selection strategy. As proof of principle we have used mir-130a and its validated target MAFB to test this strategy. Identification of MAFB and five additional targets and their subsequent confirmation as mir-130a targets by western blot analysis and knockdown experiments validates this strategy for the functional identification of miRNA targets.

Original language	English
Article number	e75
Journal	Nucleic Acids Research
Volume	40
Issue number	10
Early online date	9 Feb 2012
DOIs	https://doi.org/10.1093/nar/gks145
Publication status	Published - May 2012

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title = "A functional assay for microRNA target identification and validation",

abstract = "MicroRNAs (miRNA) are a class of small RNA molecules that regulate numerous critical cellular processes and bind to partially complementary sequences resulting in down-regulation of their target genes. Due to the incomplete homology of the miRNA to its target site identification of miRNA target genes is difficult and currently based on computational algorithms predicting large numbers of potential targets for a given miRNA. To enable the identification of biologically relevant miRNA targets, we describe a novel functional assay based on a 3'-UTR-enriched library and a positive/negative selection strategy. As proof of principle we have used mir-130a and its validated target MAFB to test this strategy. Identification of MAFB and five additional targets and their subsequent confirmation as mir-130a targets by western blot analysis and knockdown experiments validates this strategy for the functional identification of miRNA targets.",

author = "Joop Gaken and Mohamedali, {Azim M} and Jie Jiang and Farooq Malik and Doris Stangl and Smith, {Alexander E} and Constantinos Chronis and Kulasekararaj, {Austin G} and Thomas, {N Shaun B} and Farzin Farzaneh and Mahvash Tavassoli and Mufti, {Ghulam J}",

year = "2012",

month = may,

doi = "10.1093/nar/gks145",

language = "English",

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T1 - A functional assay for microRNA target identification and validation

AU - Gaken, Joop

AU - Mohamedali, Azim M

AU - Jiang, Jie

AU - Malik, Farooq

AU - Stangl, Doris

AU - Smith, Alexander E

AU - Chronis, Constantinos

AU - Kulasekararaj, Austin G

AU - Thomas, N Shaun B

AU - Farzaneh, Farzin

AU - Tavassoli, Mahvash

AU - Mufti, Ghulam J

PY - 2012/5

Y1 - 2012/5

N2 - MicroRNAs (miRNA) are a class of small RNA molecules that regulate numerous critical cellular processes and bind to partially complementary sequences resulting in down-regulation of their target genes. Due to the incomplete homology of the miRNA to its target site identification of miRNA target genes is difficult and currently based on computational algorithms predicting large numbers of potential targets for a given miRNA. To enable the identification of biologically relevant miRNA targets, we describe a novel functional assay based on a 3'-UTR-enriched library and a positive/negative selection strategy. As proof of principle we have used mir-130a and its validated target MAFB to test this strategy. Identification of MAFB and five additional targets and their subsequent confirmation as mir-130a targets by western blot analysis and knockdown experiments validates this strategy for the functional identification of miRNA targets.

AB - MicroRNAs (miRNA) are a class of small RNA molecules that regulate numerous critical cellular processes and bind to partially complementary sequences resulting in down-regulation of their target genes. Due to the incomplete homology of the miRNA to its target site identification of miRNA target genes is difficult and currently based on computational algorithms predicting large numbers of potential targets for a given miRNA. To enable the identification of biologically relevant miRNA targets, we describe a novel functional assay based on a 3'-UTR-enriched library and a positive/negative selection strategy. As proof of principle we have used mir-130a and its validated target MAFB to test this strategy. Identification of MAFB and five additional targets and their subsequent confirmation as mir-130a targets by western blot analysis and knockdown experiments validates this strategy for the functional identification of miRNA targets.

U2 - 10.1093/nar/gks145

DO - 10.1093/nar/gks145

M3 - Article

C2 - 22323518

SN - 1362-4962

VL - 40

JO - Nucleic Acids Research

JF - Nucleic Acids Research

IS - 10

M1 - e75

ER -

A functional assay for microRNA target identification and validation

Abstract

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