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Nucleic Acids Research Advance Access originally published online on October 11, 2007
Nucleic Acids Research 2007 35(19):e132; doi:10.1093/nar/gkm830
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Nucleic Acids Research, 2007, Vol. 35, No. 19 e132
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Methods Online

A novel method for poly(A) fractionation reveals a large population of mRNAs with a short poly(A) tail in mammalian cells

Hedda A. Meijer1, Martin Bushell1, Kirsti Hill1, Timothy W. Gant3, Anne E. Willis1, Peter Jones2 and Cornelia H. de Moor1,*

1RNA Biology Group, Centre for Biomolecular Sciences, School of Pharmacy, University of Nottingham, Nottingham, NG7 2RD, 2Centre for Biochemistry and Cell Biology, School of Biomedical Sciences, Queen's Medical Centre, Nottingham NG7 2UH and 3MRC Toxicology Unit, Hodgkin Building, University of Leicester, Leicester, LE1 9HN, UK

* To whom correspondence should be addressed. Tel: +44 0 1159515041; Fax: +44 0 1159515078; Email: cornelia.demoor{at}nottingham.ac.uk

Received July 10, 2007. Revised September 7, 2007. Accepted September 21, 2007.

The length of the poly(A) tail of an mRNA plays an important role in translational efficiency, mRNA stability and mRNA degradation. Regulated polyadenylation and deadenylation of specific mRNAs is involved in oogenesis, embryonic development, spermatogenesis, cell cycle progression and synaptic plasticity. Here we report a new technique to analyse the length of poly(A) tails and to separate a mixed population of mRNAs into fractions dependent on the length of their poly(A) tails. The method can be performed on crude lysate or total RNA, is fast, highly reproducible and minor changes in poly(A) tail length distribution are easily detected. We validated the method by analysing mRNAs known to undergo cytoplasmic polyadenylation during Xenopus laevis oocyte maturation. We then separated RNA from NIH3T3 cells into two fractions with short and long poly(A) tails and compared them by microarray analysis. In combination with the validation experiments, the results indicate that ~25% of the expressed genes have a poly(A) tail of less than 30 residues in a significant percentage of their transcripts.


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