Plant Molecular Biology

, Volume 68, Issue 4, pp 479–491

Homologous recombination properties of OsRad51, a recombinase from rice

Authors

  • Chittela Rajanikant
    • Plant Biochemistry Section, Molecular Biology DivisionBhabha Atomic Research Center
  • Michael Melzer
    • Leibniz Institute of Plant Genetics and Crop Plant Research (IPK)
  • Basuthkar J. Rao
    • Department of Biological SciencesTata Institute of Fundamental Research, Colaba
    • Plant Biochemistry Section, Molecular Biology DivisionBhabha Atomic Research Center
Article

DOI: 10.1007/s11103-008-9385-6

Cite this article as:
Rajanikant, C., Melzer, M., Rao, B.J. et al. Plant Mol Biol (2008) 68: 479. doi:10.1007/s11103-008-9385-6

Abstract

cDNA corresponding to OsRad51 protein was isolated from cDNA library of rice flowers (Oryzasativa, Indica cultivar group) and cloned in to pET28a expression vector. The protein was over expressed in E. coli BL21 (DE3) and purified. Purified OsRad51 could bind single and double stranded DNA, however it showed higher affinity for single stranded DNA. Transmission Electron Microscopy (TEM) studies of OsRad51–DNA complexes showed that this protein formed ring like structures and bound DNA forming filaments. OsRad51 protein promoted renaturation of complementary single strands in to duplex DNA molecules and also showed ATPase activity, which was stimulated by single strand DNA. Fluorescence resonance energy transfer (FRET) assays revealed that OsRad51 promoted homology dependent renaturation as well as strand exchange reactions. Renaturation activity was ATP dependent; however strand exchange activity was ATP independent. This is the first report on in vitro characterization of Rad51 protein from crop plants.

Keywords

ATPaseDNA bindingFRETRenaturationStrand exchangeTransmission Electron Microscopy

Abbreviations

FRET

Fluorescence resonance energy transfer

IPTG

Isopropyl thio-galactoside

MALDI-TOF

Matrix assisted laser desorption and time of flight

Ni-CAM

Nickel chelating affinity matrix

ss

Single-stranded

ds

Double-stranded

RF

Replicative form

TEM

Transmission Electron Microscopy

Supplementary material

11103_2008_9385_MOESM1_ESM.doc (312 kb)
DOC 309 kb

Copyright information

© Springer Science+Business Media B.V. 2008