An Efficient LCM-Based Method for Tissue Specific Expression Analysis of Genes and miRNAs View Full Text


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Article Info

DATE

2016-04

AUTHORS

Vibhav Gautam, Archita Singh, Sharmila Singh, Ananda K Sarkar

ABSTRACT

Laser Capture Microdissection (LCM) is a powerful tool to isolate and study gene expression pattern of desired and less accessible cells or tissues from a heterogeneous population. Existing LCM-based methods fail to obtain high quality RNA including small RNAs from small microdissected plant tissue and therefore, are not suitable for miRNA expression studies. Here, we describe an efficient and cost-effective method to obtain both high quality RNA and miRNAs from LCM-derived embryonic root apical meristematic tissue, which is difficult to access. We have significantly modified and improved the tissue fixation, processing, sectioning and RNA isolation steps and minimized the use of kits. Isolated RNA was checked for quality with bioanalyzer and used for gene expression studies. We have confirmed the presence of 19-24 nucleotide long mature miRNAs using modified stem-loop RT-PCR. This modified LCM-based method is suitable for tissue specific expression analysis of both genes and small RNAs (miRNAs). More... »

PAGES

21577

References to SciGraph publications

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  • 2015-10-13. Coevolution Pattern and Functional Conservation or Divergence of miR167s and their targets across Diverse Plant Species in SCIENTIFIC REPORTS
  • 2007-12. Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs in PLANT METHODS
  • 2010-06-28. Fluorescence-Activated Cell Sorting in Plant Developmental Biology in PLANT DEVELOPMENTAL BIOLOGY
  • 2015-04. Laser Assisted Microdissection, an Efficient Technique to Understand Tissue Specific Gene Expression Patterns and Functional Genomics in Plants in MOLECULAR BIOTECHNOLOGY
  • 2010-11. A method for obtaining high quality RNA from paraffin sections of plant tissues by laser microdissection in JOURNAL OF PLANT RESEARCH
  • 2005-12. Laser Capture Microdissection (LCM) and Expression Analyses of Glycine max (Soybean) Syncytium Containing Root Regions Formed by the Plant Pathogen Heterodera glycines (Soybean Cyst Nematode) in PLANT MOLECULAR BIOLOGY
  • 2007-04. Conserved factors regulate signalling in Arabidopsis thaliana shoot and root stem cell organizers in NATURE
  • Identifiers

    URI

    http://scigraph.springernature.com/pub.10.1038/srep21577

    DOI

    http://dx.doi.org/10.1038/srep21577

    DIMENSIONS

    https://app.dimensions.ai/details/publication/pub.1011750804

    PUBMED

    https://www.ncbi.nlm.nih.gov/pubmed/26861910


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