title: Comparison of the MpEF1α and CaMV35 promoters for application in Marchantia polymorpha overexpression studies.
abstract: Constitutive promoters are essential tools for analyses of gene functions by transgenic approaches. For overexpression and silencing studies of genes, a ubiquitous and strong expression of genes under investigation as well as selection markers is preferred. For future applications in the emerging basal plant model system Marchantia polymorpha, a liverwort, activities of the viral 35S cauliflower mosaic virus promoter and the endogenous elongation factor 1α (MpEF1α) promoter were analyzed. Expression of the reporter gene β-glucuronidase (GUS), driven by the CaMV35 and MpEF1α promoters, was compared throughout plant development. Significant differences were observed between the two promoter activities. The CaMV35 promoter yields a weak reporter gene expression in the meristematic zones but drives a strong expression in the thallus. The MpEF1α promoter causes a strong meristematic GUS expression and is more active in female sexual tissues. Overall, the MpEF1α promoter seems to be the better option for obtaining a strong and ubiquitous transgene expression. Furthermore, a whole mount in situ hybridization protocol for Marchantia was established. Analysis of MpEF1α mRNA transcript in intact, whole tissues showed an expression pattern that is overall similar to the pattern of the GUS reporter gene expression driven by the MpEF1α promoter, including strong expression in meristematic zones. The whole mount technique reported here can be used to determine the mRNA expression in intact gemmae and archegonia, and has the potential to be applied for screening large numbers of transgenic plants, for instance to identify knock-down mutants.
authors.affiliation: Department of Botany, University of Osnabrück, Barbarastraße 11, 49069, Osnabrück, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: Althoff
authors.forename: Sarah
authors.initials: S
authors.lastname: Kopischke
authors.forename: Oliver
authors.initials: O
authors.lastname: Zobell
authors.forename: Kentaro
authors.initials: K
authors.lastname: Ide
authors.forename: Kimitsune
authors.initials: K
authors.lastname: Ishizaki
authors.forename: Takayuki
authors.initials: T
authors.lastname: Kohchi
authors.forename: Sabine
authors.initials: S
authors.lastname: Zachgo
mesh_terms: D017796:Caulimovirus / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant / Q000235:genetics*; D018014:Gene Transfer Techniques; D017343:Genes, Plant / Q000235:genetics*; D017403:In Situ Hybridization; D047008:Marchantia / Q000235:genetics / Q000378:metabolism*; D018519:Meristem / Q000378:metabolism; D020648:Peptide Elongation Factor 1 / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*
chemical_list: D017931:DNA Primers; D020648:Peptide Elongation Factor 1
references.citation: J Exp Bot. 2012 Jun;63(11):4263-73
references.pmid: 22523343
references.citation: Plant Cell. 1991 Jun;3(6):583-91
references.pmid: 1841719
references.citation: Nat Protoc. 2006;1(4):1939-46
references.pmid: 17487180
references.citation: Proc Natl Acad Sci U S A. 2006 Oct 17;103(42):15511-6
references.pmid: 17030812
references.citation: Genes Dev. 2002 Jul 1;16(13):1610-5
references.pmid: 12101120
references.citation: Biosci Biotechnol Biochem. 2013;77(1):167-72
references.pmid: 23291762
references.citation: Plant J. 2000 Sep;23(5):697-702
references.pmid: 10972895
references.citation: Theor Appl Genet. 1991 May;81(5):581-8
references.pmid: 24221371
references.citation: Plant J. 2000 Nov;24(3):421-8
references.pmid: 11069714
references.citation: Proc Natl Acad Sci U S A. 2007 Apr 10;104(15):6472-7
references.pmid: 17395720
references.citation: Annu Rev Phytopathol. 1996;34:299-323
references.pmid: 15012545
references.citation: Nucleic Acids Res. 1984 Nov 26;12(22):8711-21
references.pmid: 6095209
references.citation: BMC Biotechnol. 2004 Jul 07;4:13
references.pmid: 15239842
references.citation: J Biosci Bioeng. 2007 Jul;104(1):34-41
references.pmid: 17697981
references.citation: Mol Gen Genet. 1993 Apr;238(3):428-36
references.pmid: 8492811
references.citation: Nature. 1985 Feb 28-Mar 6;313(6005):810-2
references.pmid: 3974711
references.citation: Plant Cell Physiol. 2008 Jul;49(7):1084-91
references.pmid: 18535011
references.citation: Nucleic Acids Res. 1985 Jul 11;13(13):4777-88
references.pmid: 4022773
references.citation: Proc Natl Acad Sci U S A. 2007 Oct 16;104(42):16714-9
references.pmid: 17940055
references.citation: Science. 1996 Aug 16;273(5277):948-50
references.pmid: 8688077
references.citation: Proc Natl Acad Sci U S A. 2001 Jul 31;98(16):9454-9
references.pmid: 11481501
references.citation: J Mol Biol. 1992 Jan 5;223(1):1-7
references.pmid: 1731062
references.citation: Nat Genet. 2006 May;38(5):594-7
references.pmid: 16642024
references.citation: Planta. 2004 Mar;218(5):693-703
references.pmid: 14625773
references.citation: Plant Mol Biol. 1992 Apr;18(6):1083-9
references.pmid: 1600144
references.citation: Sci Rep. 2013;3:1532
references.pmid: 23524944