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mRNA Interactome Capture from Plant Protoplasts
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Biochemistry
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JoVE Journal Biochemistry
mRNA Interactome Capture from Plant Protoplasts

mRNA Interactome Capture from Plant Protoplasts

DOI:
10.3791/56011-v

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12:29 min

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July 28, 2017

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, , ,
1Department of Biology,KU Leuven

Chapters

  • 00:05Title
  • 01:15In Vivo mRNA-protein Crosslinking by UV Irradiation
  • 02:38Protoplast Lysis Under Denaturing Conditions
  • 03:59mRNP Pull-down and Purification by Oligo-d(T)25 Beads
  • 08:14mRBP Concentration and Protein Assays
  • 10:04Results: Capture of UV-crosslinked mRNA-bound Proteins by Oligo-d(T)25 Beads
  • 11:53Conclusion

Summary

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Here, we present an interactome capture protocol applied to Arabidopsis thaliana leaf mesophyll protoplasts. This method critically relies on in vivo UV crosslinking and allows for the isolation and identification of plant mRNA-binding proteins from a physiological environment.

Tags

Keywords: MRNA InteractomeMRNA-bound ProteomePlant ProtoplastsPost-transcriptional Gene RegulationUV CrosslinkingArabidopsis ThalianaLeaf Mesophyll ProtoplastsMMG SolutionCentrifugation

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