Protocol for the simultaneous isolation of DNA, RNA, and miRNA from a single archived Kaposi sarcoma biopsy

• Published in: STAR protocols 2024-10, Vol.5 (4), p.103365, Article 103365
• Main Authors: Scholte, Larissa L.S., Browne, Justin, Nolan, David J., St. John, Peyton, Tracy, Katherine, Thur, Rafaela S., Li, Ghangzhao, Lamers, Susanna L., Bracci, Paige, McGrath, Michael S., Bethony, Jeffrey M.
• Format: Article
• Language: English
• Subjects: Cancer; Gene Expression; Molecular Biology; Protocol; Sequencing
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2024.103365

Kaposi sarcoma (KS) punch biopsies present unique challenges for extracting nucleic acids, which can be exacerbated by their long-term stabilization in RNAlater. Here, we present a protocol for simultaneously isolating DNA, RNA, and miRNA from a single KS punch biopsy. We detail the steps for preparing reagents and supplies, disrupting KS tissue using manual and mechanical methods, isolating DNA and total RNA, evaluating nucleic acid quality, and storing nucleic acids long-term. [Display omitted] •Instructions for tissue disruption of challenging/recalcitrant samples•Protocol for the simultaneous isolation of DNA and total RNA•Optimized steps for achieving higher and more consistent nucleic acid yields•Quality control strategy for assessing DNA/RNA concentration and integrity Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Kaposi sarcoma (KS) punch biopsies present unique challenges for extracting nucleic acids, which can be exacerbated by their long-term stabilization in RNAlater. Here, we present a protocol for simultaneously isolating DNA, RNA, and miRNA from a single KS punch biopsy. We detail the steps for preparing reagents and supplies, disrupting KS tissue using manual and mechanical methods, isolating DNA and total RNA, evaluating nucleic acid quality, and storing nucleic acids long-term.