Methods Mol Biol 732:25–38Ĭhomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. López-Gomollón S (2011) Detecting sRNAs by northern blotting. Nicolas FE, Moxon S, de Haro JP et al (2010) Endogenous short RNAs generated by dicer 2 and RNA-dependent RNA polymerase 1 regulate mRNAs in the basal fungus Mucor circinelloides. Nicolas FE, Pais H, Schwach F et al (2011) mRNA expression profiling reveals conserved and non-conserved miR-140 targets. Mohorianu I, Schwach F, Jing R et al (2011) Profiling of short RNAs during fleshy fruit development reveals stage-specific sRNAome expression patterns. Lopez-Gomollon S, Beckers M, Rathjen T et al (2014) Global discovery and characterization of small non-coding RNAs in marine microalgae. Mol Plant 8:261–275Īhmed A, Ward NJ, Moxon S et al (2015) A database of microRNA expression patterns in Xenopus laevis. Martinho C, Confraria A, Elias CA et al (2015) Dissection of miRNA pathways using arabidopsis mesophyll protoplasts. Genes Dev 19:517–529Ĭastel SE, Martienssen RA (2013) RNA interference in the nucleus: roles for small RNAs in transcription, epigenetics and beyond. Tomari Y, Zamore PD (2005) Perspective: machines for RNAi. Lee Y, Ahn C, Han J et al (2003) The nuclear RNase III Drosha initiates microRNA processing. Yu Y, Zhang Y, Chen X, Chen Y (2019) Plant noncoding RNAs: hidden players in development and stress responses. Suggestions and nuances obtained from experience are included as Notes. The protocol is described to prepare most of the reagents needed in the lab, but also timesaving commercial reagent alternatives are included. ![]() This chapter describes the protocols for radioactive and non-radioactive sRNA Northern blot analysis, which includes RNA extraction, polyacrylamide gel electrophoresis, membrane transfer, hybridisation and detection of sRNA using oligonucleotide probes. Northern blotting is a widely used technique to study sRNAs because it is quantitative, relatively inexpensive, and readily available for most laboratories. The abundance and specific spatio-temporal expression of many sRNAs, especially miRNAs, is relevant for their biological function. sRNAs can be classified according to their origin and mode of action into different classes such as: microRNAs (miRNAs), small interfering RNAs (siRNAs) and PIWI-interacting RNAs (piRNAs). ![]() Small RNAs (sRNAs) are key regulators of transcriptomes and proteomes of organisms through their sequence-specific interaction with complementary RNA targets.
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