By Sweta Rani (eds.)
This quantity comprises finished descriptions of miRNA biogenesis and their function within the improvement and development of varied human illnesses. the 1st few chapters of MicroRNA Profiling: tools and Protocols discuss the results of over-expressing and repressing of a objective miRNA and their results on telephone viability and proliferation. the following couple of chapters discover the protocols for overall RNA isolation from cells and cell-derived product together with formalin fastened paraffin embedded tissue and plant tissue. the previous couple of chapters talk about isolation and characterization of exosomes from medium conditioned through cellphone traces, serum, and plasma specimens. This ebook additionally comprises discussions of numerous software program instruments, resembling miRandola, PicTar, DIANA, and miRWalk. Written within the hugely profitable Methods in Molecular Biology sequence structure, chapters contain introductions to their respective themes, lists of the required fabrics and reagents, step by step, effectively reproducible laboratory protocols, and pointers on troubleshooting and averting identified pitfalls.
Comprehensive and state of the art, MicroRNA Profiling: tools and Protocols is a precious source for someone attracted to the sector of Micro RNAs.
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Additional resources for MicroRNA Profiling: Methods and Protocols
Add a further 500 μL Buffer RPE to the spin column. Centrifuge for 2 min at 8000 × g to wash the spin column membrane. Discard the collection tube with flow through. 5 RNA Elution 1. Place the RNeasy MinElute spin column in a new 2 mL collection tube. Open the lid of the spin column and centrifuge at full speed for 5 min. Discard flow through (see Note 4). 2. 5 mL collection tube (supplied with kit). 3. Add 14–30 μL pre-heated (37–42 °C) Nuclease-free water directly to the centre of the spin column membrane.
Centrifuge at 20,000 × g for 10 min at 4 °C, and discard the supernatant. 11. Wash the precipitant twice with 80 % ethanol, and dissolve in 50 μL TE buffer. 12. For electrophoresis, denature 3 mg LMW RNA of each sample and load onto urea (8 %) 15 % polyacrylamide gels. After electrophoresis, stain the gels with 1 mg/L ethidium bromide for 30 min and visualized under ultraviolet light. 4 An Economical Method for Low Molecular Weight RNA Isolation 1. Pre-warm the extraction buffer to 65 °C and add 200 μL of β mercaptoethanol to the tube.
Utensils that would not resist this temperature can be cleaned with 1 % hydrogen peroxide followed by rinsing with DEPC-water. All the reagents used should be prepared using DEPC-treated autoclaved distilled water. 1 M NaOH and 100 mM EDTA) for 2 h and rinsed well with DEPC-treated water. Make sure all the components used for agarose gel electrophoresis are RNase-free. Plant tissue harvesting for RNA isolation should be expeditious, snap freeze in liquid nitrogen, and should not be thawed until processing.