, Volume 249, Issue 6, pp 2015–2020 | Cite as

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

  • Swati Verma
  • Vibhav Gautam
  • Ananda K. SarkarEmail author
Short Communication


Main conclusion

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.


Bud meristem Gene expression Laser capture microdissection Malus MicroRNAs RNA isolation Small RNA Stem-loop RT-PCR Tree 



Diethyl pyrocarbonate


Laser capture microdissection


Tert-butyl alcohol



SV acknowledges Department of Science and Technology-Science and Engineering Research Board (DST-SERB), Govt. of India, for National-Postdoctoral Fellowship (N-PDF file 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.

Compliance with ethical standards

Conflict of interest

The authors declare no competing interests.

Supplementary material

425_2019_3127_MOESM1_ESM.pptx (2 mb)
Fig. S1 LCM system settings for laser-based excision of meristem tissue from apple bud sections (PPTX 2028 kb)
425_2019_3127_MOESM2_ESM.xlsx (16 kb)
Supplementary material 2 (XLSX 15 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Swati Verma
    • 1
  • Vibhav Gautam
    • 1
    • 2
  • Ananda K. Sarkar
    • 1
    Email author
  1. 1.National Institute of Plant Genome ResearchNew DelhiIndia
  2. 2.Centre of Experimental Medicine and Surgery, Institute of Medical SciencesBanaras Hindu UniversityVaranasiIndia

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