In Vitro Silencing of lncRNA Expression Using siRNAs
Recent advances in sequencing technologies have uncovered the existence of thousands of long noncoding RNAs (lncRNAs) with dysregulated expression in cancer. As a result, there is burgeoning interest in understanding their function and biological significance in both homeostasis and disease. RNA interference (RNAi) enables sequence-specific gene silencing and can, in principle, be employed to silence virtually any gene. However, when applied to lncRNAs, it is important to consider current limitations in their annotation and current principles regarding lncRNA regulation and function when assessing their phenotype in cancer cell lines. In this chapter we describe the analysis of lncRNA splicing variant expression, including subcellular localization, transfection of siRNAs in cancer cell lines, and validation of gene silencing by quantitative PCR and single molecule in situ hybridization. All protocols can be performed in a laboratory with essential equipment for cell culture, molecular biology, and imaging.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2021 |
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| Erschienen: |
2021 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:2348 |
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| Enthalten in: |
Methods in molecular biology (Clifton, N.J.) - 2348(2021) vom: 23., Seite 141-156 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Thijssen, Meike S [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 01.09.2021 Date Revised 06.10.2021 published: Print Citation Status MEDLINE |
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| doi: |
10.1007/978-1-0716-1581-2_9 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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NLM327080892 |
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| 520 | |a Recent advances in sequencing technologies have uncovered the existence of thousands of long noncoding RNAs (lncRNAs) with dysregulated expression in cancer. As a result, there is burgeoning interest in understanding their function and biological significance in both homeostasis and disease. RNA interference (RNAi) enables sequence-specific gene silencing and can, in principle, be employed to silence virtually any gene. However, when applied to lncRNAs, it is important to consider current limitations in their annotation and current principles regarding lncRNA regulation and function when assessing their phenotype in cancer cell lines. In this chapter we describe the analysis of lncRNA splicing variant expression, including subcellular localization, transfection of siRNAs in cancer cell lines, and validation of gene silencing by quantitative PCR and single molecule in situ hybridization. All protocols can be performed in a laboratory with essential equipment for cell culture, molecular biology, and imaging | ||
| 650 | 4 | |a Journal Article | |
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| 650 | 4 | |a Alternative splicing | |
| 650 | 4 | |a Cancer | |
| 650 | 4 | |a In situ hybridization | |
| 650 | 4 | |a Long noncoding RNAs | |
| 650 | 4 | |a Small interfering RNAs | |
| 650 | 4 | |a Subcellular localization | |
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| 650 | 7 | |a RNA, Small Interfering |2 NLM | |
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| 700 | 1 | |a Arnes, Luis |e verfasserin |4 aut | |
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