PMID- 23770918
OWN - NLM
STAT- MEDLINE
DCOM- 20140131
LR  - 20211021
IS  - 1940-087X (Electronic)
IS  - 1940-087X (Linking)
IP  - 76
DP  - 2013 Jun 7
TI  - Efficient and rapid isolation of early-stage embryos from Arabidopsis thaliana 
      seeds.
LID - 10.3791/50371 [doi]
LID - 50371
AB  - In flowering plants, the embryo develops within a nourishing tissue - the 
      endosperm - surrounded by the maternal seed integuments (or seed coat). As a 
      consequence, the isolation of plant embryos at early stages (1 cell to globular 
      stage) is technically challenging due to their relative inaccessibility. 
      Efficient manual dissection at early stages is strongly impaired by the small 
      size of young Arabidopsis seeds and the adhesiveness of the embryo to the 
      surrounding tissues. Here, we describe a method that allows the efficient 
      isolation of young Arabidopsis embryos, yielding up to 40 embryos in 1 hr to 4 
      hr, depending on the downstream application. Embryos are released into isolation 
      buffer by slightly crushing 250-750 seeds with a plastic pestle in an Eppendorf 
      tube. A glass microcapillary attached to either a standard laboratory pipette 
      (via a rubber tube) or a hydraulically controlled microinjector is used to 
      collect embryos from droplets placed on a multi-well slide on an inverted light 
      microscope. The technical skills required are simple and easily transferable, and 
      the basic setup does not require costly equipment. Collected embryos are suitable 
      for a variety of downstream applications such as RT-PCR, RNA sequencing, DNA 
      methylation analyses, fluorescence in situ hybridization (FISH), immunostaining, 
      and reporter gene assays.
FAU - Raissig, Michael T
AU  - Raissig MT
AD  - Institute of Plant Biology and Zurich-Basel Plant Science Center, University of 
      Zurich.
FAU - Gagliardini, Valeria
AU  - Gagliardini V
FAU - Jaenisch, Johan
AU  - Jaenisch J
FAU - Grossniklaus, Ueli
AU  - Grossniklaus U
FAU - Baroux, Celia
AU  - Baroux C
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Video-Audio Media
DEP - 20130607
PL  - United States
TA  - J Vis Exp
JT  - Journal of visualized experiments : JoVE
JID - 101313252
SB  - IM
MH  - Arabidopsis/*embryology/genetics
MH  - DNA Methylation
MH  - Genes, Reporter
MH  - In Situ Hybridization, Fluorescence
MH  - Seeds/embryology/genetics
PMC - PMC3735039
EDAT- 2013/06/19 06:00
MHDA- 2014/02/01 06:00
PMCR- 2015/06/07
CRDT- 2013/06/18 06:00
PHST- 2013/06/18 06:00 [entrez]
PHST- 2013/06/19 06:00 [pubmed]
PHST- 2014/02/01 06:00 [medline]
PHST- 2015/06/07 00:00 [pmc-release]
AID - 50371 [pii]
AID - 10.3791/50371 [doi]
PST - epublish
SO  - J Vis Exp. 2013 Jun 7;(76):50371. doi: 10.3791/50371.