Abstract
MicroRNAs have emerged as – important posttranscriptional regulators of gene expression. Small RNA cloning is a powerful method to identify new microRNAs (miRNAs) and to profile miRNA expression. In addition, it reveals end heterogeneity that may be important in miRNA function. Here, we describe a protocol that is optimized to clone small RNAs from limited amounts of starting material. This is often the case for studying miRNAs in a highly purified population of immune cells or other primary cell types with limited numbers. The small RNAs cloned with this protocol will have a 5′-PO4 and 3′-OH group, typical features of miRNAs, so majority of the cloned small RNAs will be miRNAs.
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Lee RC, Feinbaum RL, Ambros V. (1993) The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 75:843–54.
Lagos-Quintana M, Rauhut R, Lendeckel W, Tuschl T. (2001) Identification of novel genes coding for small expressed RNAs. Science 294:853–8.
Lau NC, Lim LP, Weinstein EG, Bartel DP. (2001) An abundant class of tiny RNAs with probable regulatory roles in Caenorhabditis elegans. Science 294:858–62.
Lee RC, Ambros V. (2001) An extensive class of small RNAs in Caenorhabditis elegans. Science 294:862–4.
Tuschl T, Zamore PD, Lehmann R, Bartel DP, Sharp PA. (1999) Targeted mRNA degradation by double-stranded RNA in vitro. Genes Dev 13:3191–7.
Neilson JR, Zheng GX, Burge CB, Sharp PA. (2007) Dynamic regulation of miRNA expression in ordered stages of cellular development. Genes Dev 21:578–89.
Wu H, Neilson JR, Kumar P, et al. (2007) miRNA profiling of naive, effector and memory CD8 T cells. PLoS One 2:e1020.
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Wu, H., Neilson, J., Manjunath, N. (2010). MicroRNA Cloning from Cells of the Immune System. In: Monticelli, S. (eds) MicroRNAs and the Immune System. Methods in Molecular Biology, vol 667. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-811-9_5
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DOI: https://doi.org/10.1007/978-1-60761-811-9_5
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Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-60761-810-2
Online ISBN: 978-1-60761-811-9
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