RNA Isolation From Meloidogyne Spp. Galls
根结线虫(Meloidogyne Spp)瘤中RNA的分离
发布: 2013年09月05日第3卷第17期 DOI: 10.21769/BioProtoc.879 浏览次数: 9647
评审: Tie Liu
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参见作者原研究论文
The authors used this protocol in:
mar 2013
Abstract
We describe an efficient method to obtain a sufficient quantity of RNA from nematode-induced galls with a high quality and integrity, proved to be appropriate for transcriptomic analysis, i.e. real time PCR, microarray hybridization or second generation sequencing. This protocol is efficient for small quantities of galls (organs with high protein and sugar contents). The protocol allows obtaining an RNA yield of 5-15 μg total RNA from 250-300 hand dissected galls at 3 days post infection (dpi) (Figure 1). It was proved particularly for Arabidopsis and tomato.
Keywords: Galls (瘿)
Meloidogyne (根结线虫)
Efficient RNA extraction (高效RNA提取)
Transcriptomics (transcriptomics)
High quality RNa (高质量RNA)
Figure 1. 3 dpi Arabidopsis gall from M. javanica. Blue lines indicate the collected material, galls with a small portion of roots (that allows easy handling), frozen and processed for RNA extraction.
Materials and Reagents
- Arabidopsis infected by M. javanica (3 dpi or 7 dpi)
- TRI Reagent (Molecular Research Centre, MRC, catalog number: TR-118 )
- Chloroform (Scharlau, catalog number: CL02031000 )
- Isopropanol (Merck KGaA, catalog number: 0 9634 )
- Sodium citrate (Sigma-Aldrich, catalog number: s4641 )
- Sodium chloride (Duchefa, catalog number: S0520 )
- NaOH (Duchefa, catalog number: s0523 )
- Ethanol (Merck KGaA, catalog number: 100986 )
- Diethylpyrocarbonate (DEPC) (Sigma-Aldrich, catalog number: D5758 )
- RNase-free water
- Acetone (Thermo Fisher Scientific, catalog number: A/0600/17 )
- Liquid nitrogen
- RNeasy Mini Kit (QIAGEN, catalog number: 74104 ))
- RNase-Free DNase Set (QIAGEN, catalog number: 79254 )
- High salt precipitation solution (see Recipes)
Equipment
- Centrifuges
- Microfuge
- 1.5 ml Eppendorf® tubes
- Porcelain mortar and pestles, textured surface on bowl interior, 60 mm diameter
- Fume hood
Procedure
文章信息
版权信息
© 2013 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
García, A., Barcala, M., Cabrera, J., Fenoll, C. and Escobar, C. (2013). RNA Isolation From Meloidogyne Spp. Galls. Bio-protocol 3(17): e879. DOI: 10.21769/BioProtoc.879.
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分类
植物科学 > 植物生理学 > 胞内共生
分子生物学 > RNA > RNA 提取
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