RNA Extraction from RNase-Rich Senescing Leaf Samples

Susheng Gan
2012 Bio-protocol  
Isolation of intact, full-length high quality RNAs is essential for RNA sequencing, reverse transcription PCR analysis of gene expression as well as RNA gel blot analysis. This simple yet easy protocol is developed to meet this need; in addition to regular samples, this protocol is especially good for isolating RNAs from RNase-rich samples such as senescing leaves and ripening fruits (from which RNAs isolated using standard method are generally degraded to certain degree). The total RNA yield
more » ... ries from 900 μg total RNA/g non-senescing leaves to 200 μg total RNA/g senescent leaves. Materials and Reagents 1. Guanidinium thiocyanate (Thermo Fisher Scientific, catalog number: BP221) 2. NaCitrate 3. N-Lauroylsarcosine (Sigma-Aldrich, catalog number: L9150) 4. β-mercaptoethanol (β-ME) 5. Glacial HAc 6. NaOH 7. Phenol (J.T.Baker ® , catalog number: 2859) 8. Ethylenediaminetetraacetic acid (EDTA) 9. Sodium dodecyl sulfate (SDS) (Sigma-Aldrich, catalog number: L5750)
doi:10.21769/bioprotoc.248 fatcat:cxtzfoqsaffytidxsjwsxqcany