Abstract
T-DNA insertional based strategies have been successfully employed to identify genes and promoters in plants. A promoter trap mutant line anth85, carrying T-DNA insertion in the 5′-UTR region of the Atprx18 (At2G24800) gene was found to exhibit GUS gene expression characteristically in anthers and cotyledons of Arabidopsis. The homozygous mutant, anth85 lacking Atprx18 transcript did not exhibit any discernable phenotypic aberration. Transcript analysis in wild type Arabidopsis reveals abundance of the Atprx18 transcript in the inflorescence. 5′ RACE analysis of the transcript reveals ‘C’ nucleotide lying 58 bp upstream from ATG as the Transcription Start Site (TSS). Deletion analysis of the upstream sequences of the Atprx18 gene led to identification of a 507 bp (−507/−1) promoter sequence sufficient to drive the reporter gene expression preferentially in anthers.
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Abbreviations
- T-DNA:
-
Transferred DNA
- 5′-RACE:
-
Random amplification of cDNA ends for 5′ end
- TSS:
-
Transcription start site
- UTR:
-
Untranslated region
- PCR:
-
Polymerase chain reaction
- RT-PCR:
-
Reverse transcriptase polymerase chain reaction
References
Borg M, Brownfield L, Twell D (2009) Male gametophyte development: a molecular perspective. J Exp Bot 60:1465–1478
Clough SJ, Bent AF (1998) Floral dip a simplified method for agrobacteium mediated transformation of Arabidopsis thaliana. Plant J 16:735–743
Cosio C, Dunand C (2009) Specific functions of individual class III peroxidases genes. J Exp Bot 60:391–408
Cosio C, Dunand C (2010) Transcriptome analysis of various flower and silique development stages indicates a set of class III peroxidase genes potentially involved in pod shattering in Arabidopsis thaliana. BMC Genomics 11:528
Hortensteiner S (2009) Stay-green regulates chlorophyll and chlorophyll-binding protein degradation during senescence. Trends Plant Sci 14:155–162
Jefferson RA (1987) Assaying chimeric genes in plants: the GUS gene fusion system. Plant Mol Biol Rep 5:387–405
Koltunow AM, Truettner J, Cox KH, Wallroth M, Goldberg RB (1990) Different temporal and spatial gene expression patterns occur during anther development. Plant Cell 2:1201–1224
McCormick S (1993) Male gametophyte development. Plant Cell 5:1265–1275
Meinke DW, Meinke LK, Showalter TC, Schissel AM, Mueller LA, Tzafrir I (2003) A sequence based map of Arabidopsis genes with mutant phenotypes. Plant Physiol 131:409–418
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497
Ostergaard L, Yanofsky MF (2004) Establishing gene function by mutagenesis in Arabidopsis thaliana. Plant J 39:682–696
Passardi F, Cosio C, Penel C, Dunand C (2005) Peroxidases have more functions than a Swiss army knife. Plant Cell Rep 24:255–265
Prasad AM, Sivanandan C, Resminath R, Thakare DR, Bhat SR, Srinivasan R (2005) Cloning and characterization of a pentatrichopeptide protein encoding gene (LOJ) that is specifically expressed in lateral organ junctions in Arabidopsis thaliana. Gene 353:67–79
Radhamony RN, Prasad AM, Srinivasan R (2005) T-DNA insertional mutagenesis in Arabidopsis: a tool for functional genomics. Elect J Biot 8:82–106
Saha D, Kumar V, Bhat SR, Srinivasan R (2011) Characterization of the upstream sequence of the LOJ gene leads to identification of a novel enhancer element conferring lateral organ junction-specific expression in Arabidopsis. Plant Mol Biol Rep 29:265–277
Sivanandan C, Sujatha TP, Prasad AM, Resminath R, Thakare DR, Bhat SR, Srinivasan R (2005) T-DNA tagging and characterization of a cryptic root-specific promoter in Arabidopsis. Biochem Bioph Acta 1731:202–208
Srinivasan R, Saha D (2009) Promoter trapping in plants using T-DNA mutagenesis. In: Jain SM, Brar DS (eds) Molecular techniques in crop improvement. Springer, Netherlands, pp 545–577
Tallis MJ, Lin Y, Rogers A, Zhang J, Street NR, Miglietta F, Karnosky DF, De Angelis P, Calfapietra C, Taylor G (2010) The transcriptome of Populus in elevated CO2 reveals increased anthocyanin biosynthesis during delayed autumnal senescence. New Phytologist 186:415–428
Thakare DR, Radhamony RN, Chikhale NJ, Bhat SR, Srinivasan R (2006) Molecular and histochemical Analysis of a T-DNA Tagged Mutant of Arabidopsis thaliana exhibiting anther-specific GUS expression. J Plant Biochem Biotech 15:7–13
Tongolli M, Penel C, Greppin H, Simon P (2002) Analysis and expression of the class III peroxidase large gene family in Arabidopsis thaliana. Gene 288:129–138
Wei W, Twell D, Lindsey K (1997) A novel nucleic acid helicase gene identified by promoter trapping in Arabidopsis. Plant J 11:1307–1314
Wellmer F, Riechmann JL, Ferreira MA, Meyerowitz EM (2004) Genome wide analysis of spatial gene expression in Arabidopsis flowers. Plant Cell 16:1314–1326
Winter D, Vinegar B, Nahal H, Ammar R, Wilson GV, Provart J (2007) An “electronic fluorescent pictograph” browser for exploring and analyzing large-scale biological data sets. PLoS ONE 2:e718. doi:10.1371/journal.pone.0000718
Acknowledgement
We thank National Agricultural Technology Project (NATP), ICAR and Department of Biotechnology (DBT) New Delhi for financial assistance.
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Kumar, V., Thakare, D.R., Saha, D.N. et al. Characterization of upstream sequences of the peroxidase gene, Atprx18 of Arabidopsis thaliana . J. Plant Biochem. Biotechnol. 21, 121–127 (2012). https://doi.org/10.1007/s13562-011-0068-z
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DOI: https://doi.org/10.1007/s13562-011-0068-z