@article{fdi:010063952,
  title = {{A}nefficient method for quantitative, single-cell analysis of chromatin modification and nuclear architecture in whole-mount ovules in {A}rabidopsis},
  author = {{S}he, {W}. {J}. and {G}rimanelli, {D}aniel and {B}aroux, {C}.},
  editor = {},
  language = {{ENG}},
  abstract = {{I}n flowering plants, the somatic-to-reproductive cell fate transition is marked by the specification of spore mother cells ({SMC}s) in floral organs of the adult plant. {T}he female {SMC} (megaspore mother cell, {MMC}) differentiates in the ovule primordium and undergoes meiosis. {T}he selected haploid megaspore then undergoes mitosis to form the multicellular female gametophyte, which will give rise to the gametes, the egg cell and central cell, together with accessory cells. {T}he limited accessibility of the {MMC}, meiocyte and female gametophyte inside the ovule is technically challenging for cytological and cytogenetic analyses at single cell level. {P}articularly, direct or indirect immunodetection of cellular or nuclear epitopes is impaired by poor penetration of the reagents inside the plant cell and single-cell imaging is demised by the lack of optical clarity in whole-mount tissues. {T}hus, we developed an efficient method to analyze the nuclear organization and chromatin modification at high resolution of single cell in whole-mount embedded {A}rabidopsis ovules. {I}t is based on dissection and embedding of fixed ovules in a thin layer of acrylamide gel on a microscopic slide. {T}he embedded ovules are subjected to chemical and enzymatic treatments aiming at improving tissue clarity and permeability to the immunostaining reagents. {T}hose treatments preserve cellular and chromatin organization, {DNA} and protein epitopes. {T}he samples can be used for different downstream cytological analyses, including chromatin immunostaining, fluorescence in situ hybridization ({FISH}), and {DNA} staining for heterochromatin analysis. {C}onfocal laser scanning microscopy ({CLSM}) imaging, with high resolution, followed by 3{D} reconstruction allows for quantitative measurements at single-cell resolution.},
  keywords = {{P}lant {B}iology ; {I}ssue 88 ; {A}rabidopsis thaliana ; ovule ; chromatin ; modification ; nuclear architecture ; immunostaining ; {F}luorescence in situ ; {H}ybridization ; {FISH} ; {DNA} staining ; {H}eterochromatin},
  booktitle = {},
  journal = {{J}ove-{J}ournal of {V}isualized {E}xperiments},
  numero = {88},
  pages = {e51530 [9 p.]},
  ISSN = {1940-087{X}},
  year = {2014},
  DOI = {10.3791/51530},
  URL = {https://www.documentation.ird.fr/hor/fdi:010063952},
}