Advertisement

Calpain pp 103-108 | Cite as

Use of the β-Glucuronidase (GUS) Reporter System to Localize Promoter Activities of the Endogenous Plant Calpain DEFECTIVE KERNEL1 (DEK1)

  • Zhe Liang
  • Hilde-Gunn Opsahl-SortebergEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1915)

Abstract

The DEFECTIVE KERNEL1 (DEK1) gene encodes the unique plant calpain protein, which is fundamental for cell specification, development, and growth of land plants. The β-glucuronidase (GUS) reporter gene system has been used to characterize and localize gene expression over several decades. When cloning a promoter upstream of the uidA/GUS reporter gene, you visualize when the promoter is activating expression by monitoring enzymatic activity of GUS, by detecting β-glucuronidase cleavage products. Here we describe a protocol for monitoring the DEK1 promoter activity in different tissues in Arabidopsis by GUS staining.

Key words

DEFECTIVE KERNEL1 DEK1 Calpain GUS uidA Arabidopsis 

Notes

Acknowledgments

We thank Stein Erik Lid for cloning the DEK1::uidA construct [7] and Dr. Lisha Shen for critical reading of the manuscript. This work was supported by the Norwegian Research Council and NMBU to H-G.O-S.

References

  1. 1.
    Jefferson RA, Kavanagh TA, Bevan MW (1987) GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907CrossRefGoogle Scholar
  2. 2.
    Liang Z, Demko V, Wilson RC, Johnson KA, Ahmad R, Perroud PF, Quatrano R, Zhao S, Shalchian-Tabrizi K, Otegui MS, Olsen OA, Johansen W (2013) The catalytic domain CysPc of the DEK1 calpain is functionally conserved in land plants. Plant J 75:742–754CrossRefGoogle Scholar
  3. 3.
    Lid SE, Gruis D, Jung R, Lorentzen JA, Ananiev E, Chamberlin M, Niu X, Meeley R, Nichols S, Olsen OA (2002) The defective kernel 1 (dek1) gene required for aleurone cell development in the endosperm of maize grains encodes a membrane protein of the calpain gene superfamily. Proc Natl Acad Sci U S A 99:5460–5465CrossRefGoogle Scholar
  4. 4.
    Sorimachi H, Hata S, Ono Y (2011) Impact of genetic insights into calpain biology. J Biochem 150:23–37CrossRefGoogle Scholar
  5. 5.
    Zhao S, Liang Z, Demko V, Wilson R, Johansen W, Olsen OA, Shalchian-Tabrizi K (2012) Massive expansion of the calpain gene family in unicellular eukaryotes. BMC Evol Biol 12:193CrossRefGoogle Scholar
  6. 6.
    Johansen W, Ako AE, Demko V, Perroud PF, Rensing SA, Mekhlif AK, Olsen OA (2016) The DEK1 calpain linker functions in three-dimensional body patterning in Physcomitrella patens. Plant Physiol 172:1089–1104PubMedPubMedCentralGoogle Scholar
  7. 7.
    Liang Z, Brown RC, Fletcher JC, Opsahl-Sorteberg HG (2015) Calpain-mediated positional information directs cell wall orientation to sustain plant stem cell activity, growth and development. Plant Cell Physiol 56:1855–1866CrossRefGoogle Scholar
  8. 8.
    Jefferson RA, Burgess SM, Hirsh D (1986) Beta-Glucuronidase from Escherichia coli as a gene-fusion marker. Proc Natl Acad Sci U S A 83:8447–8451CrossRefGoogle Scholar
  9. 9.
    Opsahl-Sorteberg HG, Divon HH, Nielsen PS, Kalla R, Hammond-Kosack M, Shimamoto K, Kohli A (2004) Identification of a 49-bp fragment of the HvLTP2 promoter directing aleurone cell specific expression. Gene 341:49–58CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Biological SciencesNational University of SingaporeSingaporeSingapore
  2. 2.Department of Plant Sciences, BIOVITNorwegian University of Life SciencesÅsNorway

Personalised recommendations