Protein Contamination in Purified DNA (HMW DNA Extraction (Monarch))

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Purified DNA shows poor A260/A230 ratio or visible turbidity. Solution may appear cloudy due to protein contamination, affecting downstream applications. Common causes: 1) Blood: incomplete or too-rapid erythrocyte lysis, carrying over hemoglobin beyond Proteinase K capacity 2) Blood: excess supernatant (> 20 µl) left on leukocyte pellet, overwhelming Proteinase K 3) Blood from erythrocyte-rich species (e.g., rabbit) not fully depleted during lysis 4) Tissue: material from protein phase transferred during phase separation 5) Tissue: input amount exceeding recommendations, causing high viscosity and reduced Proteinase K efficiency 6) Undissolved DNA causing turbidity and poor A260/A230, falsely interpreted as protein Solutions: 1) Follow erythrocyte lysis times carefully; do not rush through steps to ensure complete hemoglobin removal 2) Remove supernatant carefully, leaving ≤ 20 µl on leukocyte pellet to minimize protein carryover 3) For erythrocyte-rich species blood: reduce input to 200 µl instead of 500 µl 4) During tissue phase separation: do not transfer any protein phase material; leave ~10 µl of DNA phase or ~30 µl if protein phase not visible 5) Use recommended tissue input amounts to maintain optimal viscosity for Proteinase K digestion 6) If turbidity persists: ensure DNA is completely dissolved at 56°C/37°C/room temperature before interpreting as protein contamination