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Improved laser capture microdissection (LCM)‑based method for isolation of RNA, including miRNA and expression analysis in woody apple bud meristem

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Title Improved laser capture microdissection (LCM)‑based method for isolation of RNA, including miRNA and expression analysis in woody apple bud meristem
 
Creator Verma, Swati
Gautam, Vibhav
Sarkar, Ananda K.
 
Subject Bud meristem
Gene expression
Laser capture microdissection
Malus
MicroRNAs
RNA isolation
Small RNA
Stem-loop RT-PCR
Tree
 
Description Accepted date: 4 March 2019
Isolation of high-quality RNA, including miRNA, from microscopic woody apple bud meristem using laser capture microdissection-based method.

It is often challenging to study the expression of microRNAs (miRNAs) or genes in less accessible inner tissues of tree species rich in polyphenols or polysaccharides. Here, we report a laser capture microdissection (LCM)-based method for efficient and cost-effective isolation and expression analysis of miRNAs and genes in the meristem tissue of woody apple bud. The tissue fixation, processing, infiltration, and sectioning steps were optimized for LCM-based excision and subsequent RNA isolation. Further, we have confirmed that RNA isolated from LCM-derived apple bud meristem contained miRNAs and was of good quantity and quality, sufficient for downstream expression analysis.
SV acknowledges Department of Science and
Technology-Science and Engineering Research Board (DST-SERB),
Govt. of India, for National-Postdoctoral Fellowship (N-PDF fle no.
PDF/2016/002423). AKS and VG acknowledge funding from Department of Biotechnology (DBT), Govt. of India (research grant no. BT/
PR12766/BPA/118/63/2015). We acknowledge LCM facility, other
central instrument facility and internal grant of NIPGR. We thank Dr.
Mohar Singh, ICAR-NBPGR, Regional Station Shimla, India for giving us access to collect apple bud samples. We thank the lab members
(Shalini and Archita) for critical reading of the manuscript and valuable comments.
 
Date 2019-04-18T10:52:42Z
2019-04-18T10:52:42Z
2019
 
Type Article
 
Identifier Planta, 249(6): 2015-2020
1432-2048
http://223.31.159.10:8080/jspui/handle/123456789/944
https://link.springer.com/article/10.1007%2Fs00425-019-03127-0
https://doi.org/10.1007/s00425-019-03127-0
 
Language en_US
 
Format application/pdf
 
Publisher Springer Nature