Purified RNA Shows Degradation (RNA Cleanup)

Question

Accepted Answer

Bioanalyzer or gel electrophoresis reveals smearing or loss of distinct RNA bands. RNA integrity number (RIN) is significantly reduced compared to input material. Common causes: 1) RNase contamination from lab bench surfaces, gloves, or non-RNase-free plasticware 2) Use of non-disposable pipet tips or microfuge tubes that may harbor RNase activity 3) Kit components left unsealed or exposed to ambient air during processing 4) Improper post-purification storage - RNA stored at temperatures above -70°C or subjected to freeze-thaw cycles Solutions: 1) Work on a clean, RNase-free lab bench; wipe surfaces with RNase decontamination solution before use 2) Wear powder-free gloves throughout the procedure; change gloves frequently to prevent cross-contamination 3) Use only disposable RNase-free pipet tips and microfuge tubes (not provided in kit); avoid autoclaved reusable plasticware 4) Keep all kit components tightly sealed when not in use; minimize exposure time during reagent handling 5) Use purified RNA immediately in downstream applications or aliquot and store at -70°C; avoid repeated freeze-thaw cycles